AMINO ACID DERIVATIVES IN BACTERIAL METABOLISM* I ...

AMINO

ACID

I. DERIVATIVES

DERIVATIVES

OF LEUCINE, PHENYLALANINE, AND VALINEt BY

(From

IN BACTERIAL

CHARLES

H. EADES,

METABOLISM* TRYPTOPITAN,

JR.

the Department of Chemistry, School of Biological TJniversil?l of Tennessee, Memphis)

Sciences,

(Reccivcd for publication, July 11, 1950)

EXPERIMENTAL

The synthetic medium, Table I, was a modification of that employed by McMahan and Snell (12). Microorganisms used were La&bacillus arabinosus 17-5 (8014), Lactobacillus casei (7469), and Leuconostoc mesenteroidesP-60 (8042). All cultures were obtained from the American Type Culture Collection of Georgetown University, Washington, D. C., and were maintained on microassay culture agar (Difco) by semimonthly transfers. Inocula for use in the tests were 18 hour cultures grown in microinoculum broth (Difco) which were washed twice by centrifugation in sterile isotonic phosphate buffer (pH 6.8). Finally the crop of organisms was suspended in enough of the sterile buffer to give a turbidity reading of 65 in an 18 mm. culture tube in a Coleman junior spectropho* This work was supportedin part by the University of Tennessee Reservefor Researchand a grant of aminoacidsand vitamins from Merck and Company. The folic acid usedwaskindly suppliedby the Lederle LaboratoriesDivision, American CyanamidCompany. A preliminary report of these studieswas made before the AmericanSociety of I 100 y per 10 ml. of medium) of indolepyruvic acid, whereas L. mesenteroides failed completely to grow under identical conditions. Slow conversion by the organisms may occur, but inst~ability of the let0 acid, even under great precaution, may be such that, t,lle compound was destroyed before the organisms could use it. Dimct,hylpyruvic and LXket,oisocaproic acids seem IO be mom stable t,o aut oclaving t,han phenylpyruvic acid, since the activity found here corresponds to 1hat found by Hegsted (9), who added the sterile kdo acids after the medium was autoclaved. It has been shown that certain acetyldehydroamino acids are not used by the rat for growth (24, 25--28). From our experimental data it appears that the acetyldehydroamino acids tested are also not effective in supporting growth of the microorganisms employed here. It seems evident that bacterial organisms exhibit wide variations with respect to the utilization of amino acid derivatives. Each organism possesses its individual metabolic patterns, the character of which must be established by experimentation. ,icet,yl derivatives and keto analogues of amino acids already known to be ut,ilizable by higher animals and to be present in various biological systems are now shown t)o support growth of certain microorganisms but not others. It is apparent t,hat I,. mesereteroides is more specific in its amino acid requirements than L. arabinosus or I,. casei. Accordingly, it, should afford assay resuhs wit’h greamr valid

C.

H.

EADES,

151

JR.

ity. However, the availability of organisms which are known to respond in characteristically different manner to various derivatives introduces the possibility of identifying a biologically active derivative, even in the presence of the free amino acid. Unpublished data from this laboratory, involving additional organisms, indicate that the acetyl and keto derivatives of some amino acids may very well be differentiated from the amino acid and quantitatively estimated by the use of several organisms in parallel assay. SUMMARY

BIBLIOGRAPHY

1. 2. 3. 4. 5. 6. 7.

8. 9. 10. 11.

Eades, C. H., Jr., Federation Proc., 9, 166 (1950). Simmonds, S., Tatum, E. L., and Fruton, J. S., J. Biol. Chem., 170, 483 (1946). Simmonds, S., Tatum, E. L., and Fruton, J. S., J. Biol. Chem., 169, 91 (1947). Fruton, J. S., Simmonds, S., and Smith, V. A., J. Biol. Chem., 169, 267 (1947). Krehl, W. A., and Fruton, J. S., J. Biol. Chem., 173, 479 (1948). Agren, G., Acta them. Stand., 2, 611 (1948). Agren, G., Acta physiol. Stand., 13, 347 (1947). Simmonds, S., and Fruton, J. S., Science, 109, 561 (1947). Hegsted, D. M., J. BioZ. Chem., 16’7, 741 (1945). Broquist, H. P., and Snell, E. E., J. BioZ. Chem., 180, 59 (1949). Lyman, C. M., Kuiken, K. A., Blotter, L., and Hale, F., J. Biol. Chem.,167,395 (1945).

12. McMahan, J. R., and Snell, E. E., J. BioZ. Chem., 162, 83 (1944). 13. Prescott, B. A., Borek, E., Brecher, A., and Waelsch, H., J. Biol. 273 (1949).

Chem.,

181,

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As determined by the growth response (acid production) of the three microorganisms studied, the acetyl and chloroacetyl derivatives of DLleucine can replace the m-amino acid only for L. arabinosus; acetyl-nnor chloroacetyl-nn-tryptophan is utilizable only by L. casei; the keto analogues of leucine and valine are only 50 per cent as active as the respective n-amino acids for both L. arabinosus and L. casei, and are not utilized at all by L. mesenteroides; the keto analogues of phenylalanine and tryptophan and the acetyldehydro derivatives of leucine, phenylalanine, tryptophan, and valine are non-utilizable under conditions of our experiments. None of the test organisms utilized for growth any of the phenylalanine derivatives under our conventional assay procedure. However, when phenylpyruvic acid was sterilized by Seitz filtration and added aseptically to the previously autoclaved test system, it supported growth of all three organisms. Inasmuch as only one of the amino acid substitutes which we have employed will support the growth of L. mesenteroides, this organism appears more ‘specific than L. casei or L. arabinosus in assays for leucine, tryptophan, and valine.

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14. Consden, R., Gordon, A. H., Martin, A. J. P., Rosenheim, D., and Synge, R. L. M., Biochem. J., 39, 251 (1945). 15. Fischer, E., and Steingroever, J., Ann. Chem., 365, 167 (1909). 16. Doherty, D. G., Tietsman, J. E., and Bergmann, M., J. Biol. Chem., 147, 617 (1943). 17. Ramage, G. R., and Simonsen, J. L., J. Chem. Sot., 532 (1935). ~011. 2, 491 (1943). 18. Herbst, R. M., and Shemin, D., Org. Syntheses, 19. Leuchs, H., and Suzuki, U., Ber. them. Ges., 37, 3306 (1904). 20. Herbst, R. M., and Shemin, D., Org. Syntheses, ~011. 2, 1 (1943). ~011. 2, 519 (1943). 21. Herbst, R. M., and Shemin, D., Org. Syntheses, 22. du Vigneaud, V., and Sealock, R. R., J. Biol. Chem., 96, 511 (1932). 23. Abderhalden, E., and Kempe, M., Ber. them. Ges., 40,2737 (1907). 24. Cooley, S. L., and Wood, J. L., J. Biol. Chem., 186, 287 (1950). 25. Fodor, P. J., Price, V. E., and Greenstein, J. P., J. Biol. Chem., 182,467 (1950). 26. Hoberman, H. D., and Fruton, J. S., J. Biol. Chem., 182, 127 (1950). PTOC., 9, 147 (1950). 27. Armstrong, M. D., and Lewis, J. D., Federation 28. Wood, J. L., Cooley, S. L., and Kelley, I. M., J. Biol. Chem., 186, 641 (1950).

AMINO ACID DERIVATIVES IN BACTERIAL METABOLISM: I. DERIVATIVES OF LEUCINE, PHENYLALANINE, TRYPTOPHAN, AND VALINE Charles H. Eades, Jr. J. Biol. Chem. 1950, 187:147-152.

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