beta-Alanine betaine synthesis in the Plumbaginaceae. Purification and characterization of a trifunctional, S-adenosyl-L-methionine-dependent N-methyltransferase from Limonium latifolium leaves.
beta-Alanine (beta-Ala) betaine is an osmoprotective compound accumulated by most members of the highly stress-tolerant family Plumbaginaceae. Its potential role in plant tolerance to salinity and hypoxia makes its synthetic pathway an interesting target for metabolic engineering. In the Plumbaginaceae, beta-Ala betaine is synthesized by S-adenosyl-L-methionine-dependent N-methylation of beta-Ala via N-methyl beta-Ala and N,N-dimethyl beta-Ala. It was not known how many N-methyltransferases (NMTases) participate in the three N-methylations of beta-Ala. An NMTase was purified about 1,890-fold, from Limonium latifolium leaves, using a protocol consisting of polyethylene glycol precipitation, heat treatment, anion-exchange chromatography, gel filtration, native polyacrylamide gel electrophoresis, and two substrate affinity chromatography steps. The purified NMTase was trifunctional, methylating beta-Ala, N-methyl beta-Ala, and N,N-dimethyl beta-Ala. Gel filtration and sodium dodecyl sulfate-polyacrylamide gel electrophoresis analyses indicated that the native NMTase is a dimer of 43-kD subunits. The NMTase had an apparent K(m) of 45 microM S-adenosyl-l-methionine and substrate inhibition was observed above 200 microM. The apparent K(m) values for the methyl acceptor substrates were 5.3, 5.7, and 5.9 mM for beta-Ala, N-methyl beta-Ala, and N,N-dimethyl beta-Ala, respectively. The NMTase had an isoelectric point of 5.15 and was reversibly inhibited by the thiol reagent p-hydroxymercuribenzoic acid.
Rathinasabapathi
B
B
Horticultural Sciences Department, Plant Molecular and Cellular Biology Program, University of Florida, Gainesville, Florida 32611-0690, USA. brath@mail.ifas.ufl.edu
Fouad
W M
WM
Sigua
C A
CA
D019684:Magnoliopsida / Q000201:enzymology*; D008780:Methyltransferases / Q000302:isolation & purification / Q000378:metabolism*; D018515:Plant Leaves / Q000201:enzymology; D000644:Quaternary Ammonium Compounds; D015091:beta-Alanine / Q000031:analogs & derivatives / Q000096:biosynthesis*
D000644:Quaternary Ammonium Compounds; C434265:beta-alanine betaine; D015091:beta-Alanine; D008780:Methyltransferases; C434266:S-adenosyl-L-methionine-dependent N-methyltransferase