Abstract
Synovial fibroblasts were incubated in nutrient medium with either L-[U-14C]leucine or L-[4,5-3H]leucine to reveal peptides associated with hyaluronic acid. Isotope was found in hyaluronic acid extracted from culture medium by repeated density gradient ultracentrifugation, but 76-98% of the labelled hyaluronic acid fraction was recovered as oligosaccharide by Sephadex G-25 gel chromatography after digestion with protease-free staphylococcal hyaluronidase (EC 4.2.99.1). Radioactivity in acid digests of the oligosaccharide fractions (i) was volatile, (ii) separated similarly to acetate in Biogel P2 chromatography, and (iii) was identified as acetate by melting point after conjugation as a p-phenylphenacyl ester crystallized to constant specific activity. The synovial fibroblast thus possesses metabolic pathways for complete transformation of leucine to acetylCoA, which operate freely in adequate nutrient medium and are a significant source of acetyl groups in hyaluronic acid synthesis.
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