Cloning and characterization of phosphomannose isomerase from Sphingomonas chungbukensis DJ77.
Phosphomannose isomerase (PMI) catalyzes the interconversion of fructose-6-phosphate and mannose-6-phosphate in the extracellular polysaccharide (EPS) synthesis pathway. The gene encoding PMI in Sphingomonas chungbukensis DJ77 was cloned and expressed in E. coli. The pmi gene is 1,410 nucleotides long and the deduced amino acid sequence shares high homology with other bifunctional proteins that possess both PMI and GDP-mannose pyrophosphorylase (GMP) activities. The sequence analysis of PMI revealed two domains with three conserved motifs: a GMP domain at the N-terminus and a PMI domain at the C-terminus. Enzyme assays using the PMI protein confirmed its bifunctional activity. Both activities required divalent metal ions such as Co(2+), Ca(2+), Mg(2+), Ni(2+) or Zn(2+). Of these ions, Co(2+) was found to be the most effective activator of PMI. GDP-D-mannose was found to inhibit the PMI activity, suggesting feedback regulation of this pathway.
Tran
Sinh Thi
ST
Deparment of Biochemistry, Chungbuk National University, Cheongju 361-763, Korea.
Le
Dung Tien
DT
Kim
Young-Chang
YC
Shin
Malshik
M
Choi
Jung-Do
JD
D000595:Amino Acid Sequence; D001483:Base Sequence; D003001:Cloning, Molecular; D004591:Electrophoresis, Polyacrylamide Gel; D008359:Mannose-6-Phosphate Isomerase / Q000737:chemistry / Q000235:genetics* / Q000302:isolation & purification; D008969:Molecular Sequence Data; D016415:Sequence Alignment; D020578:Sphingomonas / Q000201:enzymology*
D008359:Mannose-6-Phosphate Isomerase