Cloning, sequence, and expression of a chitinase gene from a marine bacterium, Altermonas sp. strain O-7.
The gene encoding an extracellular chitinase from marine Alteromonas sp. strain O-7 was cloned in Escherichia coli JM109 by using pUC18. The chitinase produced was not secreted into the growth medium but accumulated in the periplasmic space. A chitinase-positive clone of E. coli produced two chitinases with different molecular weights from a single chitinase gene. These proteins showed almost the same enzymatic properties as the native chitinase of Alteromonas sp. strain O-7. The N-terminal sequences of the two enzymes were identical. The nucleotide sequence of the 3,394-bp SphI-HindIII fragment that included the chitinase gene was determined. A single open reading frame was found to encode a protein consisting of 820 amino acids with a molecular weight of 87,341. A putative ribosome-binding site, promoter, and signal sequence were identified. The deduced amino acid sequence of the cloned chitinase showed sequence homology with chitinases A (33.4%) and B (15.3%) from Serratia marcescens. Regardless of origin, the enzymes of the two bacteria isolated from marine and terrestrial environments had high homology, suggesting that these organisms evolved from a common ancestor.
Tsujibo
H
H
Osaka University of Pharmaceutical Sciences, Japan.
Orikoshi
H
H
Tanno
H
H
Fujimoto
K
K
Miyamoto
K
K
Imada
C
C
Okami
Y
Y
Inamori
Y
Y
D000595:Amino Acid Sequence; D001483:Base Sequence; D002688:Chitinases / Q000096:biosynthesis / Q000235:genetics* / Q000302:isolation & purification; D003001:Cloning, Molecular; D004252:DNA Mutational Analysis; D005798:Genes, Bacterial / Q000235:genetics*; D006088:Gram-Negative Aerobic Bacteria / Q000201:enzymology / Q000235:genetics*; D008386:Marine Biology; D008969:Molecular Sequence Data; D016366:Open Reading Frames; D011994:Recombinant Proteins / Q000096:biosynthesis / Q000235:genetics / Q000302:isolation & purification; D012045:Regulatory Sequences, Nucleic Acid / Q000235:genetics; D015183:Restriction Mapping; D017386:Sequence Homology, Amino Acid
D011994:Recombinant Proteins; D002688:Chitinases