SU717062A1 - Alpha-aminophosphonic acids for selective inhibiting of enzymes and their preparation method - Google Patents

Description

the reaction is the interaction of xyma aldehyde with anhydrous phosphorus & thystatic acid at a temperature of 25-35 ° C in an inert gas atmosphere.

Example. (Aminoisobutylphosphonist islota.

To 2.64 g (0.04 mol) of anhydrous hypophosphorous acid rijpH, with good stirring, in an argon atmosphere, add 20 pounds of 1.74 g (0.02 mol) of bainy ya oil of aldehyde, maintaining the temperature of the reaction mixture at 25-35 ° C. Permix for 1 hour (pre-refactin self-heating of the reaction mixture) and leave for 24 hours at. Equal water is then added and applied to a column (15.22.9 cm) with Dowex sulfonic resin .50 8, (100 2 O Of mesh mesh form). The column is rinsed first with water, dBtrrrrrrrrrrrrrrrrrrrn in Shtu-Tabepblokeia at 2-7 nm and neutral reactions, and then. aqueous ammonia solution1. , the final reaction with a hydronic is combined, evaporated to dryness in a vacuum, and 0.65 g is obtained.

(24%) -aminoisobutylphosphonic acid, so pl. 205-210.C (isoprpan-water).

Found,% C 35.50 H 8.98, P 22, N -.

CAHjxO. NP

Calculated,% t С 35.0) H 8.76 / hr R 22.60; N 10.20

Rf - 0.49, (upstream chromatography on Silufol UFV plates; isopropanol: ammonia: water (7: 1: 2); ninhydrin, ammonium molybdatrm) -0.92 (electrophoresis on FN-18 paper, buffer 0.05 M sodium acetate, pH 4.1, voltage 4000 volts, expressed by ninhydrin, ammonium molybdate). In bromine oxidations in a hydrochloric acid solution, the only product is d. -aminoisobutylphosphonic acid 1.

 . In a similar way, other tk-amino acid phosphonic acids obtained by this analysis and physics and chemical characteristics, of which 25 are presented in; tab. one,

CH, and. cjHaO PN Calculated,% СН, ёСЯ, СН, гГО, 91

i, - -L.,.; lv; "rV COOHCH, CHi -. , -. .

, 95 Found: C 29, la, H 6.15; R 18.70; 8.63 C4H,,.

-G - - - - calculated from 28.75 J H 5.99 R 18, 52; 3.40 (k-Aminophosphonist ACIDS are considered as a new class of analogues .a1 in a digest of acids. Replacement of the carboxyl group of the amino acid W cFyArHyTsHI of the other elements of the structure is poorly related to the next ones. synth .- ... .., ...-.- - - ..JJ.chAigl-. - .. Tazy, key enzymes of protein biosynthesis in the cell. Thanks to this, aminophosphonic acids, and n

30,165-0,19,168. C 22.60; H 7.35; R 28.41) 12.81 YAID6%: s2y, 67; H 7.35; R.18,10,8,12 40 201- 0.52 C4.HiiOiPNS - "--- 204% C 28.40 uH 7.10-, P l8.3S; 8.29 7; 43 P 28; 27 12, .5b- 20 208- 0.35 substrate amino acids are calculated, are selective and effective inhibitors of these enzymes. as shown by yppytay, conducting B1 on the purified ttfienapaTax valyl- and methionyl-tRNA synthetases from E. coS-i. In this case, an amino-isobutyl phosphonyl acid is considered as a roller analog, ati.-amino-IS-methylthiopropyl phosphonist 1 kbI and tJ as an analog of methionine. Test data are presented in table 2.

Inhibition of aminoacyl-tRYAK-synthetases d, -amino phosphonous acids

RCIKNII, P (0) (OH) H

R (CHj) jCHR CHjSCHjCHj

As can be seen from the table. The 2 analogs of vlinyl and methionine do have a specific effect on the synthetase reaction, but have high affinity for the corresponding enzymes.

Enzymes whose cofactor is pyridoxal-5-phosphate, especially transaminases, play an important role in the conversion of amino acids. In addition, the substrate of many transaminases is glutamic acid. The analogue of the latter, the k-amino-carboxypropyl phosphonate acid, was competitive with glutamic acid as a reaction inhibitor / catalyzed aspartate-glutamate transaminase from the heart of the pig.

PRI mme R 2. Inhibition of isalyl-tRNA synthetase h-aminoisobutylphosphonic acid,

Valyl tRNA synthetase is obtained from E.coe. Valine tRNA is purified from total tRNA. coSi B. For definition. dividing the enzyme activity using ATP-N and salt ik -valine (125 curimol); P - sodium pyrophosphate 16 curie-mol) and 8 -mercaptoethanl,

Inhibition of the amino-acylation reaction is carried out at 37 ° C for 15 minutes. 50 µl (5-10 M) of an h-amino-isobutylphosphonic acid solution are added to the incubation sample just prior to the addition of the enzyme. The incubation sample contained in terms of 1 ml: 40 µmol Tris-HC (pH 7.8), 5 µmol magnesium acetate, 72 µmol chloro ammonium, 6 µmol-mercggoethanol, 2 µmol ATP, 4 nmol C -i -valine , 0.3 mg tRNA and 0.005 mg enzyme. The rate of aminoacylating tRNA is determined.

The inhibition of methionyl-tRNA- (the inthetases is carried out analogously to example 2.

Example 3. Inhibition of aspartate-glutamate-transaminae from serdt a b l and c a 2

Aminoacyl-tRNA synthetase, K ,. (M) in the aminoacylation reaction; in brackets - К „for substrates

MethionylValil3-yr. {3,

pig protein (k -amino-jc-carboxypropyl phosphonic acid.

Aspartate glutamate transaminase, (EC 2,6,1,1), (AT) is obtained from the heart. pig pigs. L-aspartic acid, L-glutamic acid and c-ketoglutaric acid are used. AT activity is determined by direct optical method, registering changes in optical density at 260 nm (absorption of the enol form of oxaloacetic acid)

0 during the enzymatic reaction: (k-ketoglutaric acid + /. Aspartic acid 1-glutamic acid + oxaloacetic acid,

Inhibition is carried out under standard conditions for determining activity, at pH 8.2, K 2 ml of 0.25 Tris-HC buffer containing a mole of oxaloacetic acid, 5-10 mol of L-glutamic acid

0 and 10; mol d -amino-carboxypropylphosphonic acid, 2 10-mol enzyme is added. A decrease in norm strength at 260 nm is measured within one minute. The activity of AT is determined similarly.

with other ratios of -U-glutamic and -amino-1-carb.oxypropylphosphonic acid.

The data obtained is used for

0 L-glutamic acid competitive inhibition and for assessing the affinity of the inhibitor for the enzyme, which was only 8-10 times worse than for the substrate, L-glutamic acid.

Claims (3)

1. k-Aminophosphonous acids 0 general formula H R-CH- R f I HE NHj Oh five where R is methyl, isopropyl, jS is methylthio. -. 771 ethyl or li-carboxyethyl, for selective inhibition of enzymes,. 2. A method for the preparation of compounds according to pL, h and aaaaaaa and c with the fact that oxime aldehyde is subjected to interaction with anhydrous phosphoric acid at a temperature of 25–35 ° C in an inert gas atmosphere. 7062. 8 Sources of information taken into account in the examination of t BoiiuHhoi J: l), C.hutchev N-Y.Khomoiov R.M., Ki "$ eeev SCHLEV & Letters, -G272 22 255-257. i $ a naereonjwChew.Bioe Jut enaction, 1970, 2, 53-55. 3. Patent IT 3506579, cl. 252-107, published. 1970.