Synthesis and degradation of 1-aminocyclopropane-1-carboxylic acid by Penicillium citrinum.
1-Aminocyclopropane-1-carboxylic acid (ACC), which is a precursor of ethylene in plants, has never been known to occur in microorganisms. We describe the synthesis of ACC by Penicillium citrinum, purification of ACC synthase [EC 4.4.1.14] and ACC deaminase [EC 4.1.99.4], and their properties. Analyses of P. citrinum culture showed occurrence of ACC in the culture broth and in the cell extract. ACC synthase was purified from cells grown in a medium containing 0.05% L-methionine and ACC deaminase was done from cells incubated in a medium containing 1% 2-aminoisobutyrate. The purified ACC synthase, with a specific activity of 327 milliunit/mg protein, showed a single band of M(r) 48,000 in SDS-polyacrylamide gel electrophoresis. The molecular mass of the native enzyme by gel filtration was 96,000 Da. The ACC synthase had the Km for S-adenosyl-L-methionine of 1.74 mM and kcat of 0.56 s-1 per monomer. The purified ACC deaminase, with a specific activity of 4.7 unit/mg protein, showed one band in SDS-polyacrylamide gel electrophoresis of M(r) 41,000. The molecular mass of the native ACC deaminase was 68,000 Da by gel filtration. The enzyme had a Km for ACC of 4.8 mM and kcat of 3.52 s-1. The presence of 7 mM Cu2+ in alkaline buffer solution was effective for increasing the stability of the ACC deaminase in the process of purification.
Jia
Y J
YJ
Faculty of Agriculture, Hokkaido University, Sapporo, Japan. yjjia@chem.agr.hokudai.ac.jp
Kakuta
Y
Y
Sugawara
M
M
Igarashi
T
T
Oki
N
N
Kisaki
M
M
Shoji
T
T
Kanetuna
Y
Y
Horita
T
T
Matsui
H
H
Honma
M
M
D000596:Amino Acids / Q000096:biosynthesis / Q000378:metabolism*; D000598:Amino Acids, Cyclic; D000818:Animals; D001711:Biotransformation; D019755:Carbon-Carbon Lyases / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism; D002848:Chromatography, DEAE-Cellulose; D002852:Chromatography, Ion Exchange; D004591:Electrophoresis, Polyacrylamide Gel; D005030:Ethylenes / Q000378:metabolism; D007700:Kinetics; D008190:Lyases / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism; D008970:Molecular Weight; D030781:Organisms, Genetically Modified; D010407:Penicillium / Q000201:enzymology / Q000235:genetics / Q000378:metabolism*; D012436:S-Adenosylmethionine / Q000378:metabolism
D000596:Amino Acids; D000598:Amino Acids, Cyclic; D005030:Ethylenes; C023863:1-aminocyclopropane-1-carboxylic acid; D012436:S-Adenosylmethionine; C036216:ethylene; C033634:1-aminocyclopropane-1-carboxylate deaminase; D008190:Lyases; D019755:Carbon-Carbon Lyases; C021773:1-aminocyclopropanecarboxylate synthase