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Basis of the Stereospecific Preference of Porcine Kidney Fibroblasts for D-2-Hydroxy-4-Methylthiobutanoic Acid as a Source of Methionine^1,2,3,

Departments of Biochemistry and Animal Science, North Carolina State University, Raleigh, NC 27695

In previous studies we have found that porcine kidney fibroblasts will grow in medium containing D-2-hydroxy-4-methylthiobutanoic acid (D-methionine hydroxy analogue, D-MHA) as the sole source of methionine but not in medium containing the L-isomer (L-MHA) alone. The fibroblasts have been found to have both D-2-hydroxy acid dehydrogenase (EC 1.1.99.6), which uses D-MHA as substrate (K_m = 6.0 mM) and L-2 hydroxy acid oxidase (EC 1.1.3.1), which uses L-MHA as substrate (K_m = 7.1 mM). These two activities should make it possible for the fibroblasts to grow on either isomer. Only one protein band with L-2-hydroxy acid oxidase activity can be detected with enzyme-specific staining of protein profiles obtained after polyacrylamide gel electrophoresis. The enzyme L-2-hydroxy acid oxidase from porcine kidney has properties that are different from the two porcine isozymes reported previously by others. Passage of DL-[¹⁴C]MHA at tracer levels into the porcine kidney fibroblasts in culture is reduced to 31% of control in the presence of 3.75 mM D-MHA, 86% of control with 3.75 mM L-MHA and 65% with 3.75 mM D-lactate but is not affected by up to 3.75 mM L-lactate. It appears that the transport specificity is the basis for the growth promotion of kidney fibroblasts by the D-isomer of MHA as opposed to L-MHA when each is used as the sole source of methionine.

KEY WORDS: • L-methionine • D- and L-methionine hydroxy analogue • D-2-hydroxy acid dehydrogenase • L-2-hydroxy acid oxidase • porcine kidney fibroblasts

¹ Paper No. 11,112 of the Journal Series of the North Carolina Agricultural Research Service, Raleigh, NC 27695-7621. The use of trade names in this publication does not imply endorsement by the North Carolina Agriculture Research Service of the product named, or criticism of similar ones not mentioned.

² This research was funded partially by a research grant from Monsanto Industrial Chemicals, St. Louis, MO.

³ This report represents a part of the Ph.D. dissertation of C. L. Schreiner, submitted to the Graduate School of North Carolina State University.

Manuscript received 4 June 1987. Revision accepted 14 March 1988.