Primary structure and chemical modification of some amino acid residues of bifunctional alginate lyase from a marine bacterium Pseudoalteromonas sp. strain no. 272.
The entire amino acid sequence of bifunctional alginate lyase from Pseudoalteromonas sp. strain No. 272 were determined by two approaches, Edman degradation of the peptides obtained from protease digestion of the enzyme protein and analysis of PCR products of the structural gene. The former resulted in incomplete amino acid sequence in the entire sequence, due to lacking of the proper peptides from the protease digestion. To compensate for this lack of sequences we applied the method of PCR of the structural gene that was initially elucidated from the primers designed from N- and C-terminal amino acid sequences of the enzyme. The results of the amino acid sequences from these two approaches showed good agreement. The enzyme consisted of 233 amino acid residues with a molecular mass of 25,549.5, including the sole W and cystine residue. The sequence homology search among the other alginate lyases from different origins indicated that they were very weakly homologous, with the exception of the sequence homology (80.3%) of Pseudoalteromonas elyakovii alginate lyase. The consensus sequence, YFKhG + Y-Q (Wong, T. Y., Preston, L. A., and Schiller, N. L. 2000. Annu. Rev. MicrobioL 54: 289-340) in the C-terminal regions was conserved. The kinetic analyses of chemical modification of some amino acid residues of the enzyme showed that W, K, and Y appeared to be important in the enzyme function.
Iwamoto
Yoshiko
Y
Division of Biochemistry, Faculty of Fisheries, Nagasaki University, Nagasaki 852-8521, Japan.
Iriyama
Kenichi
K
Osatomi
Kiyoshi
K
Oda
Tatsuya
T
Muramatsu
Tsuyoshi
T
D000595:Amino Acid Sequence; D000596:Amino Acids / Q000032:analysis / Q000737:chemistry; D001483:Base Sequence; D016384:Consensus Sequence; D017931:DNA Primers; D007700:Kinetics; D008969:Molecular Sequence Data; D016133:Polymerase Chain Reaction; D011133:Polysaccharide-Lyases / Q000737:chemistry* / Q000235:genetics*; D031161:Pseudoalteromonas / Q000201:enzymology* / Q000235:genetics; D020539:Sequence Analysis, Protein / Q000379:methods; D017386:Sequence Homology, Amino Acid
D000596:Amino Acids; D017931:DNA Primers; D011133:Polysaccharide-Lyases; C016158:poly(beta-D-mannuronate) lyase