Methylmalonic aciduria is an inborn error of metabolism characterized by neonatal or infantile ketoacidosis. Leukocytes isolated from the peripheral blood of a 1-year-old child with this disorder converted negligible quantities of propionate-3-C(14) to carbon dioxide, but oxidized succinate-1,4-C(14) normally, an indication of a block in the conversion of propionate to succinate. Parenteral administration of vitamin B(18) resulted in a reduction in methylmalonic acid excretion and an increase in propionate oxidation by leukocytes in vitro. The results suggest a mutation of methylmalonyl-CoA isomerase, a vitamin B(12), dependent enzyme which converts methylmalonyl-CoA to succinyl-CoA, and provide the first demonstration of vitamin B(12) "dependency" in man.

Guinea-pig intestinal mucosa contains a microsomal enzyme which catalyses the esterification of cholesterol. The enzyme is CoA-dependent and probably an acyl-CoA-cholesterol acyltransferase (ACAT) (E.C. 2.3.1.26). The apparent Km for the CoA in the combined enzyme reaction is 3.3 x 10(-5)M. The specific activity of cholesterol esterification is about five times higher in young suckling animals than in ordinary fed adult animals. Feeding fat and fat/cholesterol diets increases the activity two- to five-fold. Concomitantly the fat/cholesterol fed animals get a marked accumulation of free and esterified cholesterol in mucosal cells.

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Human intestinal mucosa contains acyl-CoA:cholesterol acyltransferase activity. The enzyme has been studied by using oleylcarnitine, CoA and carnitine palmitoyltransferase as an oleyl-CoA regenerating system. The enzyme was found in the particulate fraction of the cells, it had a pH optimum between 7.2 and 8.2, and was inhibited by taurocholate. The specific enzymic activity in biopsies from intestinal mucosa of normal men was found to be 3.6 +/- 1.37 nmol cholesteryl ester formed mg protein-1 h-1, an activity which can account for all cholesteryl esters in intestinal lymph. Low enzymic activity was found in biopsies from patients with small intestinal disorders. Two pancreatectomized patients had values within the normal range.

Three distinct classes of human mutations (cbl A, cbl B, and cbl C) cause defective synthesis of cobalamin (Cbl; vitamin B12) coenzymes. Cultured fibroblasts from that unique class (cbl C) deficient in the synthesis of both Cbl coenzymes, 5'-deoxyadenosylcobalamin (AdoCbl) and methylcobalamin (MeCbl), were used to explore the underlying defect. We compared the uptake of transcobalamin II (TC II)-bound cyano by cbl C cells with that of other control and mutant cell lines. Although the cbl C cells initially took up CN4[7CoJCbl normally, they were unable to retain it. To characterize this "leak" further, cell extracts were prepared following incubation and chromatographed on Sephadex G-50. After incubations of 1-2 hr, most of the CN{57CoJCbl accumulated by control cells was still bound to TC II; the remainder was free. Thereafter, an ever-increasing fraction of the labeled Cbl eluted with an intracellular cobalamin-binding protein (ICB) more than 80% of the total was so bound after 76 hr incubations. ICB had an apparent molecular weight similar to that of several Cbl "R" binders (about 120,000), but was distinguished from them by its failure to react with specific anti-"R" binder antiserum. Significantly, no ICB was detected in extracts of three different cbl C lines even after prolonged incubations, whereas its appearance in cbl A, cbl B, and mutase apoenzyme mutants was normal. We propose: that ICB is required for retention of cobalamins by cels; and that cbl C cels "leak" cobalamins and show defective synthesis of Cbl coenzymes because they lack this intracellular binder.During the past 15 years a picture of the cellular biology and biochemistry of the compound originally designated vitamin B12 and now more appropriately referred to by its chemical name, cobalamin (Cbl), has begun to emerge. Based on in mvo experiments with radioisotopically labeled Cbl in man (1-3), it has been shown that, once absorbed from the small intestine, the vitamin is transported in blood tightly bound to a specific serum protein, transcobalamin II (TC II). The subsequent delivery of Cbl to hepatocytes (3-7) and other tissue cells (8-13) is mediated by a complicated, still incompletely understood process which is initiated by the binding of the TC II-Cbl complex to specific cell surface receptors. Current evidence favors the belief that the membrane-bound TC II-Cbl complex then enters the cell intact via endocytosis, and is localized transiently in secondary ly-