Abstract
Acyl-CoA synthetase activity with various long chain fatty acid substrates was measured in microsomes from rat testes, isolated spermatids and testes of hypophysectomized adult rats, using reversed-phase high performance liquid chromatography (HPLC). The spectrophotometric HPLC method produced results comparable to those of parallel radiometric assays and was highly specific for acyl-CoA products. At optimal pH and cofactor concentrations, specific activity from whole testis was similar for 18∶1, 20∶4 and 22∶5 but somewhat lower for 16∶0 over the substrate range 0.01–3.2 mM. Activity from spermatids or from testes of hypophysectomized rats was much lower with 22∶5 than with 18∶1 or 20∶4, whereas activities with 18∶1 and 20∶4 were similar at all substrate concentrations. All substrates exhibited Michaelis-Menten type saturation kinetics and linear Lineweaver-Burke plots at lower substrate concentrations but inhibited activity at higher concentrations. Apparent values of KM for 16∶0, 18∶1 and 20∶4 were more than twice that of 22∶5, whereas both observed and calculated maximum velocities were similar for the four fatty acids. Differences in pseudokinetic parameters and differential expression of the testicular acyl-CoA synthetase activities with different fatty acids suggest the presence of multiple enzymes, at least one of which may be hormonally regulated.
This is a preview of subscription content, log in via an institution to check access.
Similar content being viewed by others
References
Coniglio, J.G. (1977) inThe Testis (Johnson, A.D., and Gomes, W.R., eds.) Vol. 4, pp. 425–449, Academic Press, New York.
Coniglio, J.G. (1977) inLipid Metabolism in Mammals (Snyder, F., ed.) Vol. 2, pp. 83–129, Plenum Press, New York.
Coniglio, J.G., Whorton, A.R., and Beckman, J.K. (1977) inFunction and Biosynthesis of Lipids (Bazan, N.G., Brenner, R.R., and Giusto, N.M., eds.) p. 575–589, Plenum Press, New York.
Grogan, W.M., Farnham, W.F., and Szopiak, B.A. (1981)Lipids 16, 401–410.
Beckman, J.K., Gray, M.E., and Coniglio, J.G. (1978)Biochim. Biophys. Acta 530, 367–374.
Beckman, J.K., and Coniglio, J.G. (1979)Lipids 14, 262–267.
Ritzen, E.M., Hansson, V., and French, F.S. (1981) inThe Testis, (Burger, H. and deKretser, D., eds.) pp. 171–194, Raven Press, New York.
Grogan, W.M., and Lam, J.W. (1982)Lipids 17, 605–611.
Grogan, W.M., and Huth, E.G. (1983)Lipids 18, 275–284.
Beckman, J.K., and Coniglio, J.G. (1980)Lipids 15, 389–394.
Tanaka, T., Hosaka, K., and Numa, S. (1981)Methods Enzymol. 71, 334–341.
Wykle, R.L., Blank, M.L., and Snyder, F. (1973)Biochim. Biophys. Acta 326, 26–33.
Chaudhary, L.R. (1982)Biochimie 64, 211–217.
Lowry, O.H., Rosebrough, N.J., Farr, A.L., and Randall, R.J. (1951)J. Biol. Chem. 193, 265–275.
Wilkinson, G.N. (1961)Biochem. J. 80, 324–332.
Grogan, W.M. (1984)Lipids 19, 341–346.
Grogan, W.M., Farnham, W.F., and Sabau, J.M. (1981)J. Histochem. Cytochem. 29, 738–746.
Carpenter, M.P. (1974)Lipids 9, 397–406.
Carpenter, M.P., and Manning, L.M. (1975)Fed. Proc. 34, 672.
Aveldano, M.I., and Sprecher, H. (1983)J. Biol. Chem. 258, 9339–9343.
Smith, P.E. (1927)J. Clin. Med. Assoc. 88, 158–161.
Smith, P.E. (1930)Am. J. Anat. 45, 205–273.
Author information
Authors and Affiliations
About this article
Cite this article
Grogan, W.M., Huth, E.G. HPLC measurement of testicular long chain acyl-CoA synthetases with different substrate specificities. Lipids 21, 11–16 (1986). https://doi.org/10.1007/BF02534295
Received:
Issue Date:
DOI: https://doi.org/10.1007/BF02534295