Defective DNA Synthesis in Human Megaloblastic Bone Marrow: Effects of Hydroxy-B12 5'-Deoxyadenosyl-B12 and Methyl-B12

Abstract

Abstract In cobalamin deficiency, inadequate DNA-thymine synthesis appears to result from decreased conversion of N5-methyltetrahydrofolic acid to tetrahydrofolic acid (THF). The N5-methyl THF conversion catalyzed by N5-methyl THF-homocysteine methyltransferase requires a cobalamin coenzyme, presumed to be methylcobalamin (methyl-B12). In support of the above, in B12-deficient marrow cultures, methyl-B12 appears to be the most effective cobalamin form to correct defective DNA-thymine synthesis. This was measured by the ability of deoxyuridine to suppress tritiated thymidine incorporation into DNA. While methyl-B12 produced complete correction of defective DNA synthesis, 5'-deoxyadenosyl cobalamin (5'-deoxyadenosyl-B12), cyanocobalamin (cyano-B12), and hydroxycobalamin (hydroxy-B12) effected only partial correction. The methyl-B12-mediated correction was blocked by methotrexate (MTX). The effect of MTX, in turn, was reversed by THF. In folate-deficient marrows, the B12 analogues did not correct defective DNA-thymine synthesis. The differential effects of hydroxy-B12 and methyl-B12 in correcting defective DNA-thymine synthesis in B12-deficient marrows suggest that the complex mechanisms for N5-methyl THF-homocysteine methyltransferase activation in Escherichia coli may not predominate in human hemopoietic tissue. Since methyl-B12 is the main component of plasma cobalamins, the critical determinant for megaloblastic maturation in B12 deficiency may be the delivery rate of methyl-B12 to marrow cells and its direct activation of N5-methyl THF-homocysteine methyltransferase.