RRC ID |
48708
|
著者 |
Hirato Y, Tokuhisa M, Tanigawa M, Ashida H, Tanaka H, Nishimura K.
|
タイトル |
Cloning and characterization of d-threonine aldolase from the green alga Chlamydomonas reinhardtii.
|
ジャーナル |
Phytochemistry
|
Abstract |
d-Threonine aldolase (DTA) catalyzes the pyridoxal 5'-phosphate (PLP)-dependent interconversion of d-threonine and glycine plus acetaldehyde. The enzyme is a powerful tool for the stereospecific synthesis of various β-hydroxy amino acids in synthetic organic chemistry. In this study, DTA from the green alga Chlamydomonas reinhardtii was discovered and characterized, representing the first report to describe the existence of eukaryotic DTA. DTA was overexpressed in recombinant Escherichia coli BL21 (DE3) cells; the specific activity of the enzyme in the cell-free extract was 0.8 U/mg. The recombinant enzyme was purified to homogeneity by ammonium sulfate fractionation, DEAE-Sepharose, and Mono Q column chromatographies (purified enzyme 7.0 U/mg). For the cleavage reaction, the optimal temperature and pH were 70 °C and pH 8.4, respectively. The enzyme demonstrated 90% of residual activity at 50 °C for 1 h. The enzyme catalyzed the synthesis of d- and d-allo threonine from a mixture of glycine and acetaldehyde (the diastereomer excess of d-threonine was 18%). DTA was activated by several divalent metal ions, including manganese, and was inhibited by PLP enzyme inhibitors and metalloenzyme inhibitors.
|
巻・号 |
135
|
ページ |
18-23
|
公開日 |
2017-3-1
|
DOI |
10.1016/j.phytochem.2016.12.012
|
PII |
S0031-9422(16)30298-9
|
PMID |
28038776
|
MeSH |
Aldehyde-Lyases / metabolism
Amino Acid Sequence
Chlamydomonas reinhardtii / enzymology
Chlamydomonas reinhardtii / metabolism*
Cloning, Molecular
Escherichia coli / genetics
Glycine / metabolism
Glycine Hydroxymethyltransferase / metabolism*
Pyridoxal Phosphate / metabolism
Stereoisomerism
Substrate Specificity
Threonine
|
IF |
3.044
|
引用数 |
3
|
WOS 分野
|
BIOCHEMISTRY & MOLECULAR BIOLOGY
PLANT SCIENCES
|
リソース情報 |
藻類 |
NIES-2237 |