A novel pathway for the conversion of homocysteine to methionine in eukaryotes.
Activation of amino acid homocysteine was compared with that of methionine in rabbit crude liver extracts and purified multi-enzyme complex of aminoacyl-tRNA synthetases. Activation was studied by measuring the incorporation of radioactive amino acid into unlabelled trichloroacetic-acid insoluble materials in the absence of protein synthesis. Homocysteine synthetase activity was found in the crude extract and in the purified multi-enzyme complex of aminoacyl-tRNA synthetases. On a molar basis, the activation of methionine by the crude extract was five times higher than the activation of homocysteine. There was a partial loss of Hcy-tRNA synthetase activity in the purified multi-enzyme complex. Preliminary reconstitution experiments indicated a requirement for an additional factor for Hcy-tRNA synthetase activity. TLC of the amino acid released from tRNA charged with [14C]homocysteine, revealed radioactivity in homocysteine, methionine and homocysteine thiolactone, indicating a conversion of tRNA-attached homocysteine to methionine. Total tRNA was separated on a benzoylated cellulose column into a fraction enriched in initiator tRNA and a methionine-accepting, but initiator tRNA-deficient, fraction. Homocysteine-accepting activity was present only in the initiator tRNA-enriched fraction. Based on the above data we propose that homocysteine activation in reticulocyte lysates, reported previously, also occurs in liver. Activated homocysteine is attached to initiator tRNA and then converted to methionine by a methylating enzyme. In the absence of methylation, tRNA-attached homocysteine is hydrolysed to produce homocysteine thiolactone.
Antonio
C M
CM
Institute for Biochemistry and Molecular Biology, University of Bergen, Norway.
Nunes
M C
MC
Refsum
H
H
Abraham
A K
AK
D000818:Animals; D006710:Homocysteine / Q000031:analogs & derivatives / Q000032:analysis / Q000737:chemistry / Q000378:metabolism*; D006863:Hydrogen-Ion Concentration; D006868:Hydrolysis; D008099:Liver / Q000201:enzymology / Q000378:metabolism; D008715:Methionine / Q000378:metabolism*; D008718:Methionine-tRNA Ligase / Q000378:metabolism; D009097:Multienzyme Complexes / Q000378:metabolism; D012358:RNA, Transfer, Met / Q000737:chemistry / Q000378:metabolism*; D011817:Rabbits; D012756:Sheep
D009097:Multienzyme Complexes; D012358:RNA, Transfer, Met; D006710:Homocysteine; D008715:Methionine; C007957:homocysteine thiolactone; D008718:Methionine-tRNA Ligase