Cloning and characterization of a glucosyltransferase and a rhamnosyltransferase from Streptomyces sp. 139.
OBJECTIVE
Ste15 and ste22 present in the Ebosin biosynthesis gene cluster (ste) were previously shown to function in Ebosin biosynthesis and both of the protein products are predicted to be glycosyltransferases. In this study, their biochemical activities were confirmed.


RESULTS
ste15 and ste22 were cloned and expressed in Escherichia coli. With a continuous coupled spectrophotometric assay and using the purified proteins, we now demonstrated that the protein Ste15 has the ability of catalysing the transfer of glucose specifically from UDP-glucose to an Ebosin precursor that lacks glucose, the lipid carrier located in the cytoplasmic membrane of the gene ste15 disrupt mutant Streptomyces sp. 139 (ste15(-)). The protein Ste22 can catalyse the transfer of rhamnose specifically from TDP-rhamnose to an Ebosin precursor that lacks rhamnose, a lipophilic carrier in the cytoplasmic membrane of the gene ste22 disrupt mutant Streptomyces sp. 139 (ste22(-)).


CONCLUSIONS
The gene product of ste15 was identified to be a glucosyltransferase, and the protein encoded by ste22 was found to be a rhamnosyltransferase.


CONCLUSIONS
Both of two enzymes play essential roles in the formation of repeating units of sugars during Ebosin biosynthesis. These are the first glucosyltransferase and rhamnosyltransferase in the biosynthesis of a Streptomyces exopolysaccharide to be characterized.
Key Laboratory of Biotechnology of Antibiotics, Ministry of Health, Institute of Medicinal Biotechnology, Chinese Academy of Medical Sciences & Peking Union Medical College, Tian Tan, Beijing, China.
X
X
Li
L
L
Wang
L
L
Bai
C
C
Yao
Y
Y
Zhang
R
R
Zhang
Yuan
Y
Li
D000595:Amino Acid Sequence; D001426:Bacterial Proteins / Q000737:chemistry / Q000235:genetics* / Q000302:isolation & purification / Q000378:metabolism; D003001:Cloning, Molecular; D015870:Gene Expression; D016695:Glycosyltransferases / Q000737:chemistry / Q000235:genetics* / Q000302:isolation & purification / Q000378:metabolism; D008969:Molecular Sequence Data; D016415:Sequence Alignment; D013302:Streptomyces / Q000201:enzymology* / Q000235:genetics*; D013379:Substrate Specificity
D001426:Bacterial Proteins; D016695:Glycosyltransferases