JP7135561B2 - Analysis controller, analysis system and analysis method - Google Patents

Description

The present invention relates to an analysis control device, an analysis system, and an analysis method for analyzing biomolecules.

Abnormalities in genes involved in protein degradation mechanisms have been reported as factors of many diseases in living organisms. In addition, changes in cell or tissue conditions, such as diseases, may affect the activity of protein degradation mechanisms. Therefore, it is expected that it will become possible to elucidate the factors of diseases or to diagnose the presence, degree or type of disease by understanding changes in protein degradation mechanisms.

By analyzing biomolecules such as peptides (endogenous peptides) produced by protein degradation in vivo, it is possible to understand changes in protein degradation mechanisms. Some of the peptides are excreted outside the body together with body fluids such as blood and urine, so by analyzing such peptides, the presence or absence of disease, diseased sites or subtypes can be diagnosed in a minimally or non-invasive manner. There is a possibility that it can be done.

A mass spectrometer can be used to analyze biomolecules contained in a sample. For example, Patent Literature 1 describes a method for analyzing biomolecules using a mass spectrometer. On the other hand, Patent Document 1 also describes that the mass spectrum of a sample has too many peaks, making it difficult to extract useful information from the mass spectrum.

Therefore, in the method described in Patent Document 1, a mass spectrum of a sample containing biomolecules is obtained. Next, a mass-to-charge ratio range is selected based on, among peaks in the mass spectrum, peaks larger than a predetermined threshold. A fragmentation spectrum is acquired in a selected mass-to-charge ratio range. Biomolecules of the sample are analyzed based on the acquired fragmentation spectrum.

Japanese Patent Publication No. 2005-536728

A plurality of peptides with different sequences (peptide variants) produced from a common protein may exhibit different behaviors in response to changes in conditions such as diseases. Therefore, by combining peptide variants produced from a common protein, it may be possible to develop biomarkers indicative of specific diseases or variations in the degradation machinery of specific proteins.

However, in the analysis method of Patent Document 1, even if the sample contains peptide variants in an analyzable amount, they may be excluded from analysis. Therefore, more reliable analysis of peptide variants contained in samples is desired.

An object of the present invention is to provide an analysis control device, an analysis system, and an analysis method that enable more reliable analysis of peptides contained in samples.

(1) An analysis control device according to a first aspect of the invention is an analysis control device for controlling a mass spectrometer, wherein MS ^n-1 of a sample containing a peptide and generated without performing a selection operation by the mass spectrometer The mass-to-charge ratio of a peptide variant having a sequence obtained by adding or removing one or more amino acids from at least one of the N-terminus and C-terminus of a known peptide to a first spectrum acquisition unit that acquires a spectrum is the first An estimating unit that estimates one or more peptides that match within a predetermined range the mass-to-charge ratio of the peak of the detected intensity in the MS ^n-1 spectrum acquired by the spectrum acquisition unit of, and one or more peptides estimated by the estimating unit A selection unit that selects, from among the peptides , peptides having a degree of structural similarity equal to or higher than a predetermined threshold value with a known peptide used as a presumed source, and MS ⁿ spectra of the peptides selected by the selection unit. a setting unit for setting the ions of the peptide as precursor ions in the mass spectrometer so as to generate the peptide.

In this analysis controller an MS ^n-1 spectrum of a sample containing peptides and generated by the mass spectrometer is acquired. The mass-to-charge ratio of a peptide variant having a sequence in which one or more amino acids are added or removed from at least one of the N-terminus and C-terminus of a known peptide is the ratio of the detected intensity peak in the acquired MS ^n-1 spectrum. One or more peptides are deduced that match the mass-to-charge ratio within a predetermined range. Among the one or more estimated peptides , a peptide having a structural similarity with a known peptide from which the estimation is based is equal to or higher than a predetermined threshold is selected. The ions of the selected peptide are set as precursor ions in the mass spectrometer to generate an ^MSn spectrum of the peptide .

According to this configuration, even if a large number of peaks appear in the MS ^n-1 spectrum, the peptides corresponding to the large number of peaks are comprehensively set as targets for analysis based on the degree of structural similarity with known peptides . can do. This allows more reliable analysis of peptides contained in the sample.
Analysis of peptide variants also allows the development of biomarkers indicative of specific diseases or alterations in the degradation machinery of specific proteins. In addition, the sample can be enriched using a single antibody directed against the shared sequence. This makes it possible to establish a stable comparative quantification system that is not affected by pretreatment.

(2) The analysis control device includes a second spectrum acquisition unit configured by the setting unit and configured to acquire the MS ⁿ spectrum of precursor ions generated by the mass spectrometer, and the MS ⁿ acquired by the second spectrum acquisition unit. It may further include an identification unit that identifies peptides contained in the sample by analyzing the spectrum. This allows more reliable analysis of peptides contained in the sample.

(3) The estimation unit may estimate one or more peptides by excluding detection intensity peaks corresponding to identified peptides in the MS ⁿ⁻¹ spectrum acquired by the first spectrum acquisition unit. In this case, since the analysis of the identified peptides is omitted, the peptides contained in the sample can be efficiently analyzed in a short time.

( 4 ) The estimating unit may evaluate the degree of similarity of the peptide structure based on the shortness of the Levenshtein distance of the character string representing the sequence of the peptide variant. In this case, the degree of structural similarity between a peptide contained in a sample and a known peptide can be easily evaluated.

( 5 ) The mass spectrometer includes a liquid chromatography mass spectrometer, and the estimation unit evaluates the similarity of peptide structures based on the similarity of elution times of peptide variants in the liquid chromatography mass spectrometer. You may In this case, the degree of structural similarity between a peptide contained in a sample and a known peptide can be easily evaluated.

( 6 ) An analysis system according to the second invention comprises a mass spectrometer for generating an MS n-1 spectrum by performing MS ^n-1 analysis on a sample containing peptides , and an MS ^{n -1} spectrum generated by the mass spectrometer. and an analysis control device according to the first invention for setting peptide ions as precursor ions in the mass spectrometer based on ^one spectrum.

In this analysis system, a MS ^n-1 spectrum is generated by a mass spectrometer by performing MS ^n-1 analysis on a sample containing peptides . Based on the MS ^n-1 spectrum generated by the mass spectrometer, peptide ions are set in the mass spectrometer as precursor ions by the analysis controller.

According to this configuration, even if a large number of peaks appear in the MS ^n-1 spectrum, the peptides corresponding to the large number of peaks are comprehensively set as targets for analysis based on the degree of structural similarity with known peptides . can do. This allows more reliable analysis of peptides contained in the sample.

( 7 ) The analysis method according to the third invention is an analysis method using a mass spectrometer, wherein the MS ^n-1 spectrum of a sample containing a peptide and generated without performing a selection operation by the mass spectrometer is obtained. obtaining and mass-to-charge ratios of peptide variants having a sequence in which one or more amino acids are added or removed to at least one of the N-terminus and C-terminus of a known peptide in the obtained MS ^n-1 spectrum A step of estimating one or more peptides that match the mass-to-charge ratio of the detected intensity peak within a predetermined range ; selecting peptides for which the height of is greater than or equal to a predetermined threshold; and setting the ions of the peptides as precursor ions to a mass spectrometer to generate an MS ⁿ spectrum of the selected peptides . .

According to this analysis method, even when a large number of peaks appear in the MS ^n-1 spectrum, the peptides corresponding to the large number of peaks are comprehensively analyzed based on the degree of structural similarity with known peptides . can be set. This allows more reliable analysis of peptides contained in the sample.

According to the present invention, peptides contained in a sample can be analyzed more reliably.

It is a figure showing composition of an analysis system concerning one embodiment of the present invention. FIG. 2 is a diagram showing the configuration of the analysis control device of FIG. 1; FIG. 1 shows MS ¹ spectra of samples containing peptide variants crudely purified from urine. FIG. 4 is a diagram showing an example of peptide variant candidates estimated by an estimation unit; 4 is a flow chart showing an algorithm of analysis control processing performed by an analysis control program;

(1) Configuration of Analysis System Hereinafter, an analysis control device, an analysis system, and an analysis method according to embodiments of the present invention will be described in detail with reference to the drawings. FIG. 1 is a diagram showing the configuration of an analysis system according to one embodiment of the present invention. As shown in FIG. 1, analytical system 100 includes processor 10, mass spectrometer 30 and database storage 40. As shown in FIG.

The processing device 10 includes a CPU (central processing unit) 11, a RAM (random access memory) 12, a ROM (read only memory) 13, a storage unit 14, an operation unit 15, a display unit 16, and an input/output I/F (interface). 17 and bus 18 . CPU 11 , RAM 12 , ROM 13 , storage unit 14 , operation unit 15 , display unit 16 and input/output I/F 17 are connected to bus 18 . CPU 11 , RAM 12 and ROM 13 constitute analysis control device 20 .

RAM 12 is used as a work area for CPU 11 . A system program is stored in the ROM 13 . The storage unit 14 includes a storage medium such as a hard disk or semiconductor memory, and stores an analysis control program. Analysis control processing, which will be described later, is performed by the CPU 11 executing the analysis control program stored in the storage unit 14 on the RAM 12 .

The operation unit 15 is an input device such as a keyboard, mouse or touch panel. The display unit 16 is a display device such as a liquid crystal display device. The user can issue various instructions to the analysis control device 20 using the operation unit 15 . The display unit 16 can display the identification result by the analysis control device 20 . Input/output I/F 17 is connected to mass spectrometer 30 and database storage device 40 .

The mass spectrometer 30 is any mass spectrometer such as an ion trap type, a linear ion trap type, a tandem time-of-flight type, a quadrupole-time-of-flight type, a quadrupole-ion trap type, or a Fourier transform ion cyclotron resonance type. There may be. The mass spectrometer 30 generates a mass spectrum by performing mass spectrometry on a sample containing a plurality of endogenous peptides (peptide variants) with different sequences produced by protein degradation in vivo.

A mass spectrum is a spectrum showing the relationship between the mass-to-charge ratio and the detected intensity of ions generated from a sample within a predetermined range of mass-to-charge ratios. In this embodiment, the mass spectrum includes an MS ¹ spectrum in which ions are not dissociated and an MS ⁿ spectrum in which ions are dissociated (n−1) times. Here, n is an integer of 2 or more.

The database storage device 40 includes a large-capacity storage device such as a server, and pre-stores a large number of endogenous peptide sequence databases and a large number of full-length protein sequence databases. The database storage device 40 also stores the identification results of peptide variants previously performed by the analysis system 100 as an identification result database.

The database storage device 40 includes one or more storage devices, and the endogenous peptide sequence database, the full-length protein sequence database and the identification result database may be stored in separate storage devices. For example, a sequence database of endogenous peptides contained in urine (Mosaiques DB) is stored in an independent storage device.

The analysis controller 20 identifies peptide variants contained in the sample based on the mass spectrum generated by the mass spectrometer 30 and various databases stored in the database storage device 40 . The configuration of the analysis control device 20 will be described below.

(2) Configuration of Analysis Control Device FIG. 2 is a diagram showing the configuration of the analysis control device 20 in FIG. As shown in FIG. 2, the analysis control device 20 includes a spectrum acquisition section 21, an estimation section 22, a selection section 23, a setting section 24, a spectrum acquisition section 25, an identification section 26, and an update section 27 as functional sections. The functional units of the analysis control device 20 are implemented by the CPU 11 of FIG. 1 executing the analysis control program stored in the storage unit 14 . Some or all of the functional units of the analysis control device 20 may be realized by hardware such as electronic circuits. Also, spectrum acquisition section 21 and spectrum acquisition section 25 may be implemented as a common spectrum acquisition section.

The spectrum acquisition unit 21 acquires the MS ⁿ⁻¹ spectrum of the sample generated by the mass spectrometer 30 . The estimating unit 22 extracts the mass-to-charge ratio (hereinafter referred to as peak mass) at which the peak of the detected intensity appears in the MS ⁿ⁻¹ spectrum acquired by the spectrum acquiring unit 21 .

In the present embodiment, based on the identification result database stored in the database storage device 40, the mass-to-charge ratio corresponding to the previously identified peptide variant peak is excluded from peak mass extraction targets. be done. In this case, since the analysis of the identified peptide variants is omitted, the peptide variants contained in the sample can be efficiently analyzed in a short time.

In addition, the estimating unit 22 selects one or more peptides that match the peak mass within a predetermined mass-to-charge ratio range based on the endogenous peptide sequence database and the full-length protein sequence database stored in the database storage device 40. • Infer variant candidates. The predetermined mass-to-charge ratio range is, for example, 300 ppm to 400 ppm. Candidate peptide variants are estimated by adding one or more amino acids to at least one of the N-terminus and C-terminus of any known peptide variant (hereinafter referred to as a reference peptide variant) from which the peptide variant is estimated. It is done by adding or removing.

Here, when a chemical modification such as a label modification is introduced into a peptide variant, when estimating a candidate peptide variant, correction of the mass-to-charge ratio considering the change in the mass-to-charge ratio of the modified site is done. In addition, when the presence of a specific post-translational modification is presumed in advance, the mass-to-charge ratio is corrected assuming a combination of changes in the mass-to-charge ratio due to the presence or absence of the modified molecule. The presence of specific post-translational modifications is presumed in advance when peptide variants with specific modifications are enriched, or when the presence of frequent modifications such as oxidation of methionine residues is known. This includes cases where

When the estimation unit 22 estimates one peptide variant candidate, the selection unit 23 selects the peptide variant. On the other hand, when a plurality of peptide variant candidates are estimated by the estimating unit 22, the selecting unit 23 selects a peptide variant having the highest sequence similarity with the corresponding reference peptide variant among the plurality of peptide variant candidates. Choose a variant. In the present embodiment, the degree of similarity between peptide variant sequences is evaluated by the shortness of the Levenshtein distance (edit distance) of the character strings representing the peptide variant sequences.

The setting unit 24 sets the peptide variant ions selected by the selection unit 23 as precursor ions. Thereby, in the mass spectrometer 30, mass spectrometry of precursor ions set by the setting unit 24 is performed, and an MS ⁿ spectrum is generated. A spectrum acquisition unit 25 acquires the MS ⁿ spectrum generated by the mass spectrometer 30 . Multiple peaks corresponding to multiple product ions produced by the dissociation of precursor ions appear in the MS ⁿ spectrum.

The identification unit 26 identifies peptide variants contained in the sample by analyzing the MS ⁿ spectrum acquired by the spectrum acquisition unit 25 . Specifically, the sequence of a modified peptide variant is generated by a technique similar to the estimation of the candidate peptide variant described above. In addition, the peptide variants are ranked by comparing the product ion mass-to-charge ratios calculated from the generated peptide variant sequences with the mass-to-charge ratios of the product ion peaks detected in the MS ⁿ spectrum. , are identified. Peptide variants may be identified by conventional protein sequence database searching methods for proteome analysis (such as the Mascot MS/MS Ion Search method) or spectral library searching methods (SpectraST method).

After that, the identification unit 26 causes the display unit 16 to display the identification result. The update unit 27 causes the database storage device 40 to store the peptide variant identification results obtained by the identification unit 26 . As a result, the identification result database stored in the database storage device 40 is updated to include the new identification result.

In the above configuration, if the selection unit 23 cannot select a peptide variant, the setting unit 24 selects the MS ^n-1 precursor ion based on general criteria such as the peak mass in the MS ^n-1 spectrum. may be set. Here, if a modifying molecule not previously assumed in the MS ^n-1 product ion is dissociated by the MS ^n-1 analysis (particularly, if the modifying molecule dissociates preferentially over the peptide bond), the setting unit 24 can set the ion of the peptide variant as the MS ⁿ precursor ion.

(3) Analysis Control Processing FIG. 3 shows the MS ¹ spectrum of a sample containing peptide variants crudely purified from urine. The MS ¹ spectrum of FIG. 3 is generated by performing monoisotopic ion peak detection on a mass spectrum generated by mass spectrometer 30 using matrix-assisted laser desorption ionization (MALDI) techniques.

FIG. 4 is a diagram showing an example of peptide variant candidates estimated by the estimation unit 22 for a specific peak of the MS ¹ spectrum of FIG. FIG. 5 is a flow chart showing an analysis control processing algorithm performed by the analysis control program. As an example of the analysis control process, a process for identifying peptide variants crudely purified from urine will be described below with reference to FIGS. 3 to 5. FIG.

First, the spectrum acquisition unit 21 determines whether or not the MS ¹ spectrum in FIG. 3 has been acquired (step S1). If the MS ¹ spectrum is not acquired, the spectrum acquisition unit 21 waits until the MS ¹ spectrum is acquired. When the MS ¹ spectrum is acquired, the estimator 22 extracts the peak mass in the MS ¹ spectrum (step S2).

Here, the peak with the highest intensity appears at the mass-to-charge ratio of 1441.2. However, according to the identification results database stored in the database storage device 40, the peptide variant corresponding to this peak has previously been identified as a fragment of the fibrinogen α-chain. Therefore, the estimation unit 22 excludes the mass-to-charge ratio "1441.2" and extracts the peak mass. Similarly, the estimator 22 extracts peak masses by excluding mass-to-charge ratios corresponding to other identified peptide variants.

Only the analytical control process for the peptide variant corresponding to the peak with the second highest intensity of peak mass '1354.2' will be described below. The analysis control processing for the peptide variants corresponding to other peak masses is the same as the analysis control processing for the peptide variants corresponding to the peak mass of "1354.2", so the explanation is omitted.

Next, the estimation unit 22 estimates peptide variant candidates (step S3). Subsequently, the estimation unit 22 determines whether or not there is another peptide variant candidate (step S4). If there is another peptide variant candidate, the estimation unit 22 returns to step S3. Steps S3 and S4 are repeated until all peptide variant candidates are estimated. The sequence database of endogenous peptides used in the processes of steps S3 and S4 is Mosaiques DB, which is a sequence database of endogenous peptides contained in urine.

Putative peptide variant candidates are shown in FIG. In the example of FIG. 4, six peptide variant candidates, "candidate 1" to "candidate 6", are estimated in a mass range of, for example, 300 ppm from the peak mass "1354.2". FIG. 4 shows the mass-to-charge ratio, protein name, sequence of the peptide variant candidate, endogenous peptide start and end numbers, reference peptide variant Sequences and Levenshtein distances are given. In FIG. 4, "candidate 1" to "candidate 6" are listed in ascending order of Levenshtein distance.

Thereafter, the selection unit 23 determines whether or not there is one peptide variant candidate estimated in step S3 (step S5). If there is one estimated peptide variant candidate, the selection unit 23 selects the peptide variant (step S6) , and proceeds to step S8. On the other hand, if there is more than one estimated peptide variant candidate, the selection unit 23 selects the peptide variant with the highest sequence similarity to the reference peptide variant (step S7), and proceeds to step S8. move on. In this example, the "candidate 1" peptide variant with the shortest Levenshtein distance is selected.

The reference peptide variant corresponding to "Candidate 1" is a known fragment of fibrinogen α-chain, and its sequence is "DEAGSEADHEGTHS" (theoretical mass-to-charge ratio 1441.5). In addition, the peptide variant of "candidate 1" has a structure in which only one amino acid serine residue is truncated from the C-terminus of the reference peptide variant, and its sequence is "DEAGSEADHEGTH" (mass-to-charge ratio "1354.5 ”).

Next, in step S8, the setting unit 24 sets the ions of the peptide variant selected in step S6 or step S7 (mass-to-charge ratio "1354.5") as precursor ions (step S8). As a result, the precursor ions set in step S8 are subjected to mass spectrometry in the mass spectrometer 30 to generate an ^MS2 spectrum.

Subsequently, the spectrum acquisition unit 25 determines whether or not the ^MS2 spectrum generated after step S8 has been acquired (step S9). If the MS ² spectrum is not acquired, the spectrum acquisition unit 25 waits until the MS ² spectrum is acquired. When the MS ² spectrum is acquired, the identification unit 26 identifies peptide variants contained in the sample by analyzing the MS ² spectrum (step S10).

After that, the identification unit 26 causes the display unit 16 to display the identification result in step S10 (step S11). The update unit 27 also updates the identification result database stored in the database storage device 40 using the identification result obtained in step S10 (step S12), and ends the analysis control process. As a result of this analytical control treatment, the peptide variant contained in the sample was identified as a fragment of the fibrinogen α-chain with the sequence "DEAGSEADHEGTH" (mass to charge ratio "1354.5").

(4) Effect In the analysis system 100 according to the present invention, even if multiple peaks appear in the MS ^n-1 spectrum, each of the multiple peaks corresponds to each based on the sequence similarity with known peptide variants. Peptide variants can be comprehensively set as targets for analysis. This allows more reliable analysis of peptide variants contained in the sample.

Analysis of peptide variants also allows the development of biomarkers indicative of specific diseases or alterations in the degradation machinery of specific proteins. Furthermore, since the sample can be concentrated using one type of antibody whose antigen is the shared sequence, a stable comparative quantification system can be established that is not affected by pretreatment.

(5) Other Embodiments (a) In the above embodiments, the degree of similarity between the sequences of the peptide variants is evaluated by the short Levenshtein distance of the character strings representing the sequences of the peptide variants. , the invention is not limited thereto. The degree of sequence similarity of peptide variants may be evaluated by other indices. For example, when a liquid chromatography mass spectrometer (LC/MS) is used as the mass spectrometer 30 and the sample to be measured is separated for each component, the high degree of sequence similarity of the peptide variants indicates that the elution You may evaluate by the high degree of similarity of time.

Here, in LC/MS, multiple components of a sample are sequentially dropped onto different wells of a sample plate for each elution time. Therefore, the peptide variants contained in the components added to the wells corresponding to the wells to which the reference peptide variant was added were evaluated to have higher sequence similarity with the reference peptide variant than other peptide variants. may

Alternatively, a peptide variant whose elution time matches the elution time estimated by an elution time estimation program such as SSRC (Sequence Specific Retention Calculator) within a predetermined range is more closely related to the reference peptide variant than other peptide variants. may be evaluated as having high similarity. Elution times may also be normalized with an internal standard peptide. As a result, the elution time can be calculated accurately regardless of the measurement conditions.

(b) In the above embodiment, when there are a plurality of peptide variant candidates estimated by the estimation unit 22, the selection unit 23 selects the peptide variant with the highest sequence similarity to the reference peptide variant. However, the invention is not so limited. When there are a plurality of peptide variant candidates estimated by the estimation unit 22, the selection unit 23 may select a peptide variant whose sequence similarity to the reference peptide variant is equal to or higher than a threshold value.

(c) In the above embodiment, the analysis control device 20 identifies peptide variants, but the present invention is not limited to this. The analysis control device 20 may identify lipids, sugar chains, or complex molecules thereof produced or metabolized by biosynthesis or biodegradation mechanisms. It can also be expected to develop biomarkers using such lipids, sugar chains, or complex molecules thereof.
(6) Reference form
(6-1) The analysis control device according to the first reference embodiment is an analysis control device that controls a mass spectrometer, and is a sample containing biomolecules and generated by the mass ^spectrometer . Based on the first spectrum acquisition unit to acquire and the structure of a known biomolecule, a predetermined range for the mass-to-charge ratio at which the peak of the detection intensity appears in the MS ^n-1 spectrum acquired by the first spectrum acquisition unit A predetermined level of structural similarity between an estimating unit that estimates one or more biomolecules that match each other and a known biomolecule that is an estimation source among the one or more biomolecules estimated by the estimating unit and a setting unit for setting ions of the biomolecules selected by the selection unit as precursor ions in the mass spectrometer so as to generate an MSn spectrum of ^the biomolecules selected by the selection unit. and
In this analysis controller, an MS ^n-1 spectrum of a sample containing biomolecules and produced by a mass spectrometer is acquired. Based on known biomolecular structures, one or more biomolecules are deduced that match within a predetermined range the mass-to-charge ratio at which the detected intensity peak appeared in the acquired MS ⁿ⁻¹ spectrum. Among the one or more estimated biomolecules, biomolecules having a degree of structural similarity with a known biomolecule from which the estimation is made are greater than or equal to a predetermined threshold value are selected. The ions of the selected biomolecules are placed in the mass spectrometer as precursor ions to produce an MSn spectrum of ^the selected biomolecules .
According to this configuration, even if a large number of peaks appear in the MS ^n-1 spectrum, the biomolecules corresponding to the large number of peaks are comprehensively analyzed based on the degree of structural similarity with known biomolecules. can be set as Thereby, the biomolecules contained in the sample can be analyzed more reliably.
(6-2) The analysis control device includes a second spectrum acquisition unit that acquires the MS ⁿ spectrum of precursor ions set by the setting unit and generated by the mass spectrometer, and and an identification unit that identifies biomolecules contained in the sample by analyzing the MS ⁿ spectrum. Thereby, the biomolecules contained in the sample can be analyzed more reliably.
(6-3) The estimation unit estimates one or more biomolecules by excluding detection intensity peaks corresponding to identified biomolecules in the MS ⁿ⁻¹ spectrum acquired by the first spectrum acquisition unit . good too. In this case, since the analysis of the identified biomolecules is omitted, the biomolecules contained in the sample can be efficiently analyzed in a short time.
(6-4) The biomolecules contained in the sample may be peptide variants produced from a common protein. In this case, analysis of peptide variants makes it possible to develop biomarkers indicative of specific diseases or alterations in the degradation machinery of specific proteins. Alternatively, the sample can be enriched using a single antibody that targets the shared sequence. This makes it possible to establish a stable comparative quantification system that is not affected by pretreatment.
(6-5) The estimating unit may evaluate the degree of structural similarity of biomolecules based on the shortness of the Levenshtein distance of the character string representing the sequence of the peptide variant. In this case, it is possible to easily evaluate the degree of structural similarity between the biomolecules contained in the sample and known biomolecules.
(6-6) The mass spectrometer includes a liquid chromatography mass spectrometer, and the estimating unit has a high degree of similarity in the structure of the biomolecule due to the high similarity of the elution times of the peptide variants in the liquid chromatography mass spectrometer. can be evaluated. In this case, it is possible to easily evaluate the degree of structural similarity between the biomolecules contained in the sample and known biomolecules.
(6-7) An analysis system according to the second reference form includes a mass spectrometer that generates an MS ^n-1 spectrum by performing MS ^n-1 analysis on a sample containing biomolecules, and and an analysis control device according to the first embodiment for setting ions of biomolecules as precursor ions in the mass spectrometer based on the obtained MS ⁿ⁻¹ spectrum.
In this analysis system, a MS ^n-1 spectrum is generated by a mass spectrometer by performing MS ^n-1 analysis on a sample containing biomolecules . Based on the MS ⁿ⁻¹ spectrum generated by the mass spectrometer , ions of biomolecules are set in the mass spectrometer as precursor ions by the analysis controller.
According to this configuration, even if a large number of peaks appear in the MS ^n-1 spectrum, the biomolecules corresponding to the large number of peaks are comprehensively analyzed based on the degree of structural similarity with known biomolecules. can be set as Thereby, the biomolecules contained in the sample can be analyzed more reliably.
(6-8) The analysis method according to the third reference form is an analysis method using a mass spectrometer, in which an MS ^n-1 spectrum of a sample containing biomolecules and generated by the mass spectrometer is acquired. estimating one or more biomolecules that match within a predetermined range the mass-to-charge ratio at which the detected intensity peak appeared in the acquired MS ⁿ⁻¹ spectrum, based on known biomolecular structures ; a step of selecting biomolecules having a structural similarity higher than or equal to a predetermined threshold value with a known biomolecule used as an estimation source from among the one or more estimated biomolecules; setting the ions of the biomolecules as precursor ions in a mass spectrometer to generate an MS ⁿ spectrum of the biomolecules.
According to this analysis method, even if a large number of peaks appear in the MS ^n-1 spectrum, the biomolecules corresponding to the large number of peaks can be comprehensively analyzed based on the degree of structural similarity with known biomolecules. Can be set as a target. Thereby, the biomolecules contained in the sample can be analyzed more reliably.

DESCRIPTION OF SYMBOLS 10... Processing apparatus 11... CPU, 12... RAM, 13... ROM, 14... Storage part, 15... Operation part, 16... Display part, 17... Input/output I/F, 18... Bus, 20... Analysis control device, DESCRIPTION OF SYMBOLS 21, 25... Spectrum acquisition part 22... Estimation part 23... Selection part 24... Setting part 26... Identification part 27... Update part 30... Mass spectrometer 40... Database storage device 100... Analysis system

Claims (7)

An analysis controller for controlling a mass spectrometer,
a first spectrum acquisition unit that acquires an MS ^n-1 spectrum of a sample that contains a peptide and is generated without selection operation by the mass spectrometer;
The mass-to-charge ratio of a peptide variant having a sequence in which one or more amino acids are added or removed from at least one of the N-terminus and C-terminus of a known peptide is obtained by the first spectrum acquisition unit MS ^n- an estimation unit that estimates one or more peptides that match the mass-to-charge ratio of the detected intensity peak in ^one spectrum within a predetermined range;
a selection unit that selects, from among the one or more peptides estimated by the estimation unit, peptides having a degree of structural similarity with a known peptide from which the estimation is made is equal to or higher than a predetermined threshold;
and a setting unit for setting ions of the peptide selected by the selection unit to the mass spectrometer as precursor ions so as to generate an ^MSn spectrum of the peptide . a second spectrum acquisition unit that acquires an MS ⁿ spectrum of precursor ions set by the setting unit and generated by the mass spectrometer;
2. The analysis control device according to claim 1, further comprising an identification section that identifies peptides contained in the sample by analyzing the MS ⁿ spectrum acquired by said second spectrum acquisition section. 3. The estimation unit estimates one or more peptides by excluding detection intensity peaks corresponding to identified peptides in the MS ⁿ⁻¹ spectrum acquired by the first spectrum acquisition unit. Analytical control device as described. The analysis control according to any one of claims 1 to 3, wherein the estimating unit evaluates the degree of similarity of the peptide structure based on the shortness of Levenshtein distance of the character string indicating the sequence of the peptide variant. Device. The mass spectrometer includes a liquid chromatography mass spectrometer,
4. The estimating unit according to any one of claims 1 to 3, wherein the estimating unit evaluates the similarity of peptide structures based on the similarity of elution times of peptide variants in the liquid chromatography mass spectrometer. analysis controller. a mass spectrometer that generates an MS ^n-1 spectrum by performing MS ^n-1 analysis on a sample containing peptides ;
The analysis control device according to any one of claims 1 to 5 , wherein peptide ions are set in the mass spectrometer as precursor ions based on the MS ^n-1 spectrum generated by the mass spectrometer, analysis system. An analysis method using a mass spectrometer,
obtaining an MS ^n-1 spectrum of a sample generated containing peptides and without selection operations performed by the mass spectrometer;
The mass-to-charge ratio of a peptide variant having a sequence in which one or more amino acids are added or removed from at least one of the N-terminus and C-terminus of a known peptide is the ratio of the detected intensity peak in the acquired MS ^n-1 spectrum. predicting one or more peptides that match the mass-to-charge ratio within a predetermined range;
a step of selecting, from among the one or more deduced peptides , a peptide whose degree of structural similarity to a known peptide from which the deduction is based is equal to or higher than a predetermined threshold;
and setting the ions of the selected peptide as precursor ions in the mass spectrometer so as to generate an ^MSn spectrum of the peptide .

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