Aminotransferase activity and bioinformatic analysis of 1-aminocyclopropane-1-carboxylate synthase.
The mechanistic fate of pyridoxal phosphate (PLP)-dependent enzymes diverges after the quinonoid intermediate. 1-Aminocyclopropane-1-carboxylate (ACC) synthase, a member of the alpha family of PLP-dependent enzymes, is optimized to direct electrons from the quinonoid intermediate to the gamma-carbon of its substrate, S-adenosyl-L-methionine (SAM), to yield ACC and 5'-methylthioadenosine. The data presented show that this quinonoid may also accept a proton at C(4)' of the cofactor to yield alpha-keto acids and the pyridoxamine phosphate (PMP) form of the enzyme when other amino acids are presented as alternative substrates. Addition of excess pyruvate converts the PMP form of the enzyme back to the PLP form. C(alpha)-deprotonation from L-Ala is shown by NMR-monitored solvent exchange to be reversible with a rate that is less than 25-fold slower than that of deprotonation of SAM. The rate-determining step for transamination follows the formation of the quinonoid intermediate. The rate-determining step for alpha, gamma-elimination from enzyme-bound SAM is likewise shown to occur after C(alpha)-deprotonation, and the quinonoid intermediate accumulates during this reaction. BLAST searches, sequence alignments, and structural comparisons indicate that ACC synthases are evolutionarily related to the aminotransferases. In agreement with previously published reports, an absence of homology was found between the alpha and beta families of the PLP-dependent enzyme superfamily.
Feng
L
L
Department of Molecular and Cell Biology, University of California-Berkeley, Berkeley, California 94720-3206, USA.
Geck
M K
MK
Eliot
A C
AC
Kirsch
J F
JF
D000595:Amino Acid Sequence; D019295:Computational Biology / Q000379:methods; D019143:Evolution, Molecular; D008190:Lyases / Q000145:classification / Q000378:metabolism*; D008969:Molecular Sequence Data; D010940:Plant Proteins / Q000378:metabolism; D011732:Pyridoxal Phosphate; D020539:Sequence Analysis, Protein; D017386:Sequence Homology, Amino Acid; D013379:Substrate Specificity; D000637:Transaminases / Q000145:classification / Q000378:metabolism*
D010940:Plant Proteins; D011732:Pyridoxal Phosphate; D000637:Transaminases; D008190:Lyases; C021773:1-aminocyclopropanecarboxylate synthase