CN104359967A - Method for analyzing biomass spectrometry overlapped isotope outline - Google Patents
Method for analyzing biomass spectrometry overlapped isotope outline Download PDFInfo
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Abstract
The invention relates to a method for analyzing a biomass spectrum overlapped isotope outline. According to the method, based on the original first-stage and second-stage mass spectrums of the spectrum, the practical distribution intensity of overlapped isotopic peaks in isotope outlines is calculated by calculating the difference of the ideal experiment intensity of the overlapped isotopic peaks and the practical experiment intensity. Compared with the prior art, the method provided by the invention is small in analysis and resolution calculation amount of the intensity of the overlapped isotopic peaks, high in flux, high in accuracy, and applicable to efficient analysis and accurate structure identification on biomacromolecules (such as protein and saccharides) spectrums and cascade-stage spectrums.
Description
Technical field
The present invention relates to the analytic method of the overlapping isotope profile of a kind of biological mass spectrometry, relate generally to the system biology field relevant to biological mass spectrometry, comprise the technical fields such as proteomics, metabolism group, sugar group.
Background technology
Along with development and the commercialization of Soft ionization techniques (as electron spray ionisation) and High Resolntion Mass-Spectrometry analyzer (as track trap), mass spectrum is more and more for the analysis of biomacromolecule (as protein, polysaccharide).
Chinese patent CN 103389335 A discloses a kind of analytical equipment and the method for coming the large molecule primary structure of identification of organism and composition by analyzing biomacromolecule mass spectrometric data.This analytical equipment and method based on the original firsts and seconds mass spectrum of described biomacromolecule, are identified this biomacromolecule by isotopic peak mass-to-charge ratio and profile fingerprint comparison.The Initial experiments mass spectrometric data that above-mentioned analytical equipment and method directly utilize mass spectrometer to gather, the accurate mass-to-charge ratio of each Initial experiments isotopic peak in precursor ion and fragmention isotope profile and relative intensity and corresponding theoretical value are compared, be respectively used to find candidate biomacromolecule from database and utilize tandem mass spectrometry to confirm the highest one of wherein confidence level, thus biomolecule is carried out to the qualitative and quantitative analysis of high confidence level.
These macromolecular charged ions of mass-spectrometer measurement (precursor ion, first mass spectrometric) and corresponding fragment (fragmention, second order ms).The two-dimentional spectrogram that mass spectrum is transverse axis is mass-to-charge ratio, the longitudinal axis is intensity.Due to isotopic existence, all ions (comprising precursor ion and fragmention) are the isotope profile comprising several isotopic peaks in mass spectrum.Because mass spectrum transverse axis, namely mass analyzer has certain scope (as 500-2000) to the analysis of mass-to-charge ratio, and the fragment of biomacromolecule more (and each fragment generally occurs with multiple valence state), in second order ms, the isotope profile of fragmention has comparatively intensive overlap.Tradition fails effectively resolve these overlapping isotope profiles and split based on the method for " deconvoluting " (also referred to as removing isotope); " the isotope profile fingerprint comparison " of newly-developed, although the ratio partition method accuracy in algorithm is high, speed is slower.
Summary of the invention
Object of the present invention is exactly the analytic method providing the overlapping isotope profile of a kind of calculated amount is little, flux is high, accuracy is high biological mass spectrometry in order to overcome defect that above-mentioned prior art exists.
Object of the present invention can be achieved through the following technical solutions:
An analytic method for the overlapping isotope profile of biological mass spectrometry, comprises the following steps:
(1) finding out the theoretical relative intensity of overlapping ion and all isotopic peaks thereof and experiment absolute strength by testing the fingerprint comparison of isotope profile with isotope profile theoretical in database in mass spectrogram, comprising theoretical relative intensity and the actual experiment absolute strength sum of overlapping isotopic peak;
(2) using non-overlapping top in the theoretical isotope profile of the ion of each participation overlap as with reference to peak, the laboratory strength of each isotopic peak in corresponding experiment isotope profile is normalized;
(3) the gedanken experiment absolute strength of overlapping isotopic peak in each overlapping ion is calculated;
(4) strength variance of overlapping isotopic peak actual experiment absolute strength sum relative to gedanken experiment absolute strength sum is calculated;
(5) overlapping isotopic peak is calculated in each actual experiment absolute strength participated in overlapping ion.
In step (3), the gedanken experiment absolute strength of overlapping isotopic peak in each overlapping ion is obtained by following formulae discovery:
Gedanken experiment absolute strength=each theoretical relative intensity of theoretical relative intensity * reference isotopic peak actual experiment absolute strength/reference isotopic peak participating in overlapping isotopic peak of each overlapping isotopic peak.
In step (4), overlapping isotopic peak actual experiment absolute strength sum is obtained by following formulae discovery relative to the strength variance of gedanken experiment absolute strength sum:
Overlapping isotopic peak actual experiment absolute strength sum is relative to strength variance=[actual experiment absolute strength sum-(the gedanken experiment absolute strength summation of overlapping isotopic peak in each overlapping ion) of overlapping isotopic peak]/(the gedanken experiment absolute strength summation of overlapping isotopic peak in each overlapping ion) of gedanken experiment absolute strength sum.
In step (5), overlapping isotopic peak is obtained by following formulae discovery in each actual experiment absolute strength participated in overlapping ion:
Overlapping isotopic peak is at each gedanken experiment absolute strength * of actual experiment absolute strength=overlapping isotopic peak in each overlapping ion (1+ overlapping isotopic peak actual experiment absolute strength sum is relative to the strength variance of gedanken experiment absolute strength sum) participated in overlapping ion.
According to the overlapping isotopic peak calculated in each actual experiment absolute strength participated in overlapping ion, make the isotope profile fingerprint comparison figure after parsing.
The method that the present invention relates to is equally applicable to the parsing of overlapping profile information and data in other mass spectrums and any 2-D data.
Compared with prior art, analytic method of the present invention based on described mass spectrographic original firsts and seconds mass spectrum, by calculating the gedanken experiment intensity of overlapping isotopic peak and calculating the actual allocated intensity of overlapping isotopic peak in each overlapping isotope profile with the deviation of actual experiment intensity.Analytic method of the present invention to the parsing of the intensity of overlapping isotopic peak and split calculated amount little, flux is high, and accuracy is high, is applicable to biomacromolecule (as protein, carbohydrate) tandem mass spectrometry and efficiently resolves and structure precise Identification.
Accompanying drawing explanation
Figure 1A be the induction of ubiquitin protein energetic encounter dissociate fragmention y52-6+ mass-to-charge ratio in second order ms be 982.37582 and 983.388428 overlapping isotopic peak (in figure arrow mark) overlapping resolve before isotope profile fingerprint comparison figure;
Figure 1B be the induction of ubiquitin protein energetic encounter dissociate fragmention y52-6+ mass-to-charge ratio in second order ms be 982.37582 and 983.388428 overlapping isotopic peak (in figure arrow mark) overlapping resolve after isotope profile fingerprint comparison figure;
Fig. 2 A is that ubiquitin protein energetic encounter induces fragmention y61-H in the second order ms that dissociates
2o-7+ mass-to-charge ratio is the isotope profile fingerprint comparison figure before overlapping isotopic peak (in figure arrow mark) the overlapping parsing of 982.37582 and 983.388428;
Fig. 2 B is that ubiquitin protein energetic encounter induces fragmention y61-H in the second order ms that dissociates
2o-7+ mass-to-charge ratio is the isotope profile fingerprint comparison figure after overlapping isotopic peak (in figure arrow mark) the overlapping parsing of 982.37582 and 983.388428.
In figure, m/z is mass-to-charge ratio, and Abundance (a.u.) is the absolute laboratory strength of arbitrary unit; Theo. be theoretical isotope profile, Exp. is experiment isotope profile.
Embodiment
Below in conjunction with the drawings and specific embodiments, the present invention is described in detail.
Embodiment
An analytic method for the overlapping isotope profile of biological mass spectrometry, comprises the following steps:
(1) finding out the theoretical relative intensity of overlapping ion and all isotopic peaks thereof and experiment absolute strength by testing the fingerprint comparison of isotope profile with isotope profile theoretical in database in mass spectrogram, comprising theoretical relative intensity and the actual experiment absolute strength sum of overlapping isotopic peak;
(2) using non-overlapping top in the theoretical isotope profile of the ion of each participation overlap as with reference to peak, the laboratory strength of each isotopic peak in corresponding experiment isotope profile is normalized;
(3) the gedanken experiment absolute strength of overlapping isotopic peak in each overlapping ion is calculated:
Gedanken experiment absolute strength=each theoretical relative intensity of theoretical relative intensity * reference isotopic peak actual experiment absolute strength/reference isotopic peak participating in overlapping isotopic peak of each overlapping isotopic peak;
(4) strength variance of overlapping isotopic peak actual experiment absolute strength sum relative to gedanken experiment absolute strength sum is calculated:
Overlapping isotopic peak actual experiment absolute strength sum is relative to strength variance=[actual experiment absolute strength sum-(the gedanken experiment absolute strength summation of overlapping isotopic peak in each overlapping ion) of overlapping isotopic peak]/(the gedanken experiment absolute strength summation of overlapping isotopic peak in each overlapping ion) of gedanken experiment absolute strength sum;
(5) overlapping isotopic peak is calculated in each actual experiment absolute strength participated in overlapping ion:
Overlapping isotopic peak is at each gedanken experiment absolute strength * of actual experiment absolute strength=overlapping isotopic peak in each overlapping ion (1+ overlapping isotopic peak actual experiment absolute strength sum is relative to the strength variance of gedanken experiment absolute strength sum) participated in overlapping ion.
Then according to the actual experiment absolute strength of the overlapping isotopic peak calculated in each ion participating in overlap, the isotope profile fingerprint comparison figure after parsing is made.
Y52-6+ and y61-H in the second order ms that dissociates is induced below with ubiquitin protein energetic encounter
2the parsing of O-7+ two overlapping ions and be split as concrete implementation step and the result that example illustrates this analytic method, as shown in table 1 and Figure 1A ~ Fig. 2 B.The row and column of table 1 marks with arabic numeral and English alphabet respectively, and grid then represents with the combination of letter and number, as B6 represents that B arranges the 6th row.For the overlapping isotopic peak of mass-to-charge ratio at 982.385132 places, the first step calculates y52-6+ and y61-H with B7*D8/B8 and B12*D13/B13 respectively
2the gedanken experiment absolute strength of O-7+ ion, the numerical value namely in grid E3 and F3; Second step, calculates the strength variance of actual experiment absolute strength sum relative to gedanken experiment absolute strength sum, the numerical value namely in grid G3 with formula (D3-(E3+F3))/(E3+F3); 3rd step, calculates y52-6+ and y61-H respectively with formula E3* (1+G3) and F3* (1+G3)
2o-7+ ion mass-to-charge ratio is the actual experiment absolute strength of the isotopic peak of 982.385132.From table 1 arrange H and row G relative intensity deviation relatively, the error after parsing have dropped 88 and 125 percentage points before comparatively resolving respectively.Y52-6+ and y61-H
2the overlapping isotopic peak (mass-to-charge ratio 983.388428) of another one of O-7+ ion is carried out the result after resolving and splitting by above-mentioned same step and is also listed in table 1.The overlapping isotopic peak of y52-6+ ion resolves the isotope profile fingerprint comparison figure of front and back respectively at shown in Figure 1A, Figure 1B.Y61-H
2the overlapping isotopic peak of O-7+ ion resolves the isotope profile fingerprint comparison figure of front and back as shown in Fig. 2 A, Fig. 2 B.
Table 1
Table 1 only lists y52-6+ and y61-H
2the overlapping isotopic peak of O-7+ two ions and the isotopic peak for being normalized (being transformed into relative intensity) to experiment absolute strength.The computing formula of experiment relative intensity, mass-to-charge ratio deviation and relative intensity deviation is demonstrated as follows with the isotopic peak that mass-to-charge ratio is 982.37582: experiment relative intensity (E7)=D7*B8/D8, mass-to-charge ratio deviation (F7)=(C7-A7) * (1E-6)/A7, relative intensity deviation (G7)=E7-B7.
Relative intensity deviate in table 1 in H6 and H11 is overlapping parsing and the deviation before splitting.
Above-mentioned is can understand and use invention for ease of those skilled in the art to the description of embodiment.Person skilled in the art obviously easily can make various amendment to these embodiments, and General Principle described herein is applied in other embodiments and need not through performing creative labour.Therefore, the invention is not restricted to above-described embodiment, those skilled in the art, according to announcement of the present invention, do not depart from improvement that scope makes and amendment all should within protection scope of the present invention.
Claims (5)
1. an analytic method for the overlapping isotope profile of biological mass spectrometry, is characterized in that, comprise the following steps:
(1) finding out the theoretical relative intensity of overlapping ion and all isotopic peaks thereof and experiment absolute strength by testing the fingerprint comparison of isotope profile with isotope profile theoretical in database in mass spectrogram, comprising theoretical relative intensity and the actual experiment absolute strength sum of overlapping isotopic peak;
(2) using non-overlapping top in the theoretical isotope profile of the ion of each participation overlap as with reference to peak, the laboratory strength of each isotopic peak in corresponding experiment isotope profile is normalized;
(3) the gedanken experiment absolute strength of overlapping isotopic peak in each overlapping ion is calculated;
(4) strength variance of overlapping isotopic peak actual experiment absolute strength sum relative to gedanken experiment absolute strength sum is calculated;
(5) overlapping isotopic peak is calculated in each actual experiment absolute strength participated in overlapping ion.
2. the analytic method of the overlapping isotope profile of a kind of biological mass spectrometry according to claim 1, it is characterized in that, in step (3), the gedanken experiment absolute strength of overlapping isotopic peak in each overlapping ion is obtained by following formulae discovery:
Gedanken experiment absolute strength=each theoretical relative intensity of theoretical relative intensity * reference isotopic peak actual experiment absolute strength/reference isotopic peak participating in overlapping isotopic peak of each overlapping isotopic peak.
3. the analytic method of the overlapping isotope profile of a kind of biological mass spectrometry according to claim 2, it is characterized in that, in step (4), overlapping isotopic peak actual experiment absolute strength sum is obtained by following formulae discovery relative to the strength variance of gedanken experiment absolute strength sum:
Overlapping isotopic peak actual experiment absolute strength sum is relative to strength variance=[actual experiment absolute strength sum-(the gedanken experiment absolute strength summation of overlapping isotopic peak in each overlapping ion) of overlapping isotopic peak]/(the gedanken experiment absolute strength summation of overlapping isotopic peak in each overlapping ion) of gedanken experiment absolute strength sum.
4. the analytic method of the overlapping isotope profile of a kind of biological mass spectrometry according to claim 3, it is characterized in that, in step (5), overlapping isotopic peak is obtained by following formulae discovery in each actual experiment absolute strength participated in overlapping ion:
Overlapping isotopic peak is at each gedanken experiment absolute strength * of actual experiment absolute strength=overlapping isotopic peak in each overlapping ion (1+ overlapping isotopic peak actual experiment absolute strength sum is relative to the strength variance of gedanken experiment absolute strength sum) participated in overlapping ion.
5. the analytic method of the overlapping isotope profile of a kind of biological mass spectrometry according to claim 1, it is characterized in that, according to the overlapping isotopic peak calculated in each actual experiment absolute strength participated in overlapping ion, make the isotope profile fingerprint comparison figure after parsing.
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| CN111551626A (en) * | 2020-05-18 | 2020-08-18 | 苏州市汉诺生物科技有限公司 | Cascade mass spectrometry analysis method based on molecular composition and structural fingerprint identification |
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| CN106990159A (en) * | 2017-05-04 | 2017-07-28 | 同济大学 | A kind of protein quantitation methods based on complete accurate same weight diethyl mark |
| CN111551626A (en) * | 2020-05-18 | 2020-08-18 | 苏州市汉诺生物科技有限公司 | Cascade mass spectrometry analysis method based on molecular composition and structural fingerprint identification |
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