Identification of a novel gentisate 1,2-dioxygenase from Silicibacter pomeroyi.

Abstract

A 1,125-bp long ORF encoding a novel gentisate 1,2-dioxygenase with two-domain bicupins was cloned from Silicibacter pomeroyi DSS-3 and expressed in Escherichia coli. The resulting product was purified to homogeneity and partially characterized. Non-reductive SDS-PAGE and gel filtration showed that the active recombinant gentisate 1,2-dioxygenase had an estimated molecular mass of 132 kDa, and reductive SDS-PAGE indicated an approximate size of 45 kDa. The enzyme thus appears to be a homotrimeric protein. This is in contrast to the homotetrameric or dimeric protein of the gentisate 1,2-dioxygenases that have been characterized thus far. The K (m) and K (cat)/K (m) for gentisate were 12 muM and 653 x 10(4) M(-1 )s(-1); the pI was 4.6-4.8. It was optimally active at 40 degrees C and pH 8.0.

• Authors: Dongqi Liu, Tingting Zhu, Li Fan, Junming Quan, Hongchun Guo, Jinren Ni
• Journal: Biotechnology letters (Biotechnol Lett) Vol. 29 Issue 10 Pg. 1529-35 (Oct 2007) ISSN: 0141-5492 [Print] Netherlands
• PMID: 17684705 (Publication Type: Journal Article, Research Support, Non-U.S. Gov't)

Chemical References

• Recombinant Proteins
• Dioxygenases
• gentisate 1,2-dioxygenase

Topics

• Amino Acid Sequence
• Catalytic Domain
• Dioxygenases (chemistry, genetics, metabolism)
• Electrophoresis, Polyacrylamide Gel
• Escherichia coli (genetics)
• Models, Molecular
• Molecular Sequence Data
• Molecular Structure
• Open Reading Frames (genetics)
• Phylogeny
• Recombinant Proteins (chemistry, metabolism)
• Rhodobacteraceae (classification, enzymology, genetics)
• Sequence Homology, Amino Acid