Coenzyme A-synthesizing protein complex of Saccharomyces cerevisiae.

The coenzyme A-synthesizing protein complex (CoA-SPC) is a multienzyme complex of Saccharomyces cerevisiae (Bakers' yeast), which has a molecular weight in excess of 200,000 as determined by Sephadex G-200 column chromatography. This multienzyme complex, which is insoluble in the crude yeast cell lysate, has been purified 229-fold. A cellular component of the yeast cell lysate, referred to as t-Factor, with a molecular weight of 400-1000 and chloride ion are involved in the solubilization of CoA-SPC. The CoA-SPC requires L-cysteine, D-pantothenic acid and ATP as substrates. The terminal CoA-SPC-bound intermediate is dephospho-CoA, which is subsequently phosphorylated and released from the complex as CoA. The sequence of reactions for the synthesis of CoA by the CoA-SPC differs significantly from those previously proposed for other systems. It could be that the reaction sequence is unique for the yeast cell.

Regulation of fatty acid synthetase activity. The 4'-phosphopantetheine hydrolase of rat liver.

The 4'-phosphopantetheine hydrolase of rat liver, partially purified by ammonium sulfate precipitation, catalyzes the hydrolysis of the prosthetic group 4'-phosphopantetheine from the holo-fatty acid synthetase. The two products of the action of this enzyme, 4'-phosphopantetheine and apo-fatty acid synthetase, were isolated by DEAE-cellulose chromatography and by chromatography on a Sepharose epsilon-aminocaproyl pantetheine column, respectively. The resultant apo-fatty acid synthetase was quantitated by immunoprecipitation and it was also converted to the holoprotein with a crude preparation of rat liver 4'-phosphopantetheine transferase. Quantitative determination of the hydrolase reaction product, 4'-phosphopantetheine, by amino acid analysis and microbiological assays confirmed the presence of 1 mol of this compound/mol of holo-fatty acid synthetase.

Changes in the collagenolytic activity released by primary VX-2 carcinoma cultures as a function of tumor growth.

Serum-free media of minced tissue cultures of VX-2 rabbit carcinoma contained a specific collagenolytic activity capable of releasing soluble radioactive peptides from [14C]-labeled collagen fibrils. It was also capable of reducing the viscosity of acid-soluble collagen solutions by cleaving the tropocollagen (TC) molecules primarily at one site to TCA (75%) and TCB (25%) fragments. Three chromatographic fractions were separated by gel filtration: F1, (MW 85-110,000) present in larger amounts in early cultures of younger tumor tissue; F2, (MW-35-40,000) the major component with maximum production in the day 3 media of younger and advanced tumor tissues; F3, (MW 18-22,000) the minor component. Early cultures of younger tumor tissue contained a latent collagenase and were subject to trypsin activation suggesting the presence of inactive enzyme precursors or an enzyme-inhibitor complex.

Mechanism for the incorporation of S-(1,2,3,4-tetrahydro-2-hydroxy-1-naphthyl)-L-cysteine into protein.

Evidence is presented which indicates that S-(1,2,3,4-tetrahydro-2-hydroxy-1-naphthyl)-L-cysteine (THN-cysteine), formed by the reaction of 1,2-epoxy-THN with cysteine, can be incorporated into protein; The position of incorporation of THN-cysteine into protein would depend on whether the epoxide of THN reacts with cysteinyl-tRNACyS or with cysteine. In both cases, the mechanism of incorporation of THN-cysteine into protein is the same as for the natural amino acids. For example, the incorporation of THN-cysteinyl-tRNACyS is stimulated by Poly-UG, the code for tRNACyS, and would be expected to be substituted for cysteine in protein being synthesized, whereas THN-cysteine not previously esterified to tRNA is activated by the isoleucyl- and valyl-RNA synthetases, and its incorporation is stimulated by Poly-AU and Poly-UG, respectively. Consequently, in this case, THN-cysteine would substitute for isoleucine and valine during protein synthesis.