The cloning and characterization of a second alpha-amylase of A. hydrophila JMP636.
OBJECTIVE
The aim of this study was to identify, clone and characterize the second amylase of Aeromonas hydrophila JMP636, AmyB, and to compare it to AmyA.


RESULTS
The amylase activity of A. hydrophila JMP636 is encoded by multiple genes. A second genetically distinct amylase gene, amyB, has been cloned and expressed from its own promoter in Escherichia coli. AmyB is a large alpha-amylase of 668 amino acids. Outside the conserved domains of alpha-amylases there is limited sequence relationship between the two alpha-amylases of A. hydrophila JMP636 AmyA and AmyB. Significant (80%) similarity exists between amyB and an alpha-amylase of A. hydrophila strain MCC-1. Differences in either the functional properties or activity under different environmental conditions as possible explanations for multiple copies of amylases in JMP636 is less likely after an examination of several physical properties, with each of the properties being very similar for both enzymes (optimal pH and temperature, heat instability). However the reaction end products and substrate specificity did vary enough to give a possible reason for the two enzymes being present. Both enzymes were confirmed to be alpha-type amylases.


CONCLUSIONS
AmyB has been isolated, characterized and then compared to AmyA.


CONCLUSIONS
The amylase phenotype is rarely encoded by more than one enzyme within one strain, this study therefore allows the better understanding of the unusual amylase production by A. hydrophila.
Kidd
S P
SP
School of Biosciences, University of Birmingham, Edgbaston, B15 2TT, UK, Department of Microbiology and Parasitology, University of Queensland, St Lucia, QLD 4072. s.p.kidd@bham.ac.uk
Pemberton
J M
JM
D016980:Aeromonas hydrophila / Q000201:enzymology* / Q000235:genetics; D000595:Amino Acid Sequence; D001426:Bacterial Proteins; D001483:Base Sequence; D003001:Cloning, Molecular; D004926:Escherichia coli / Q000235:genetics; D015870:Gene Expression; D006026:Glycoside Hydrolases / Q000235:genetics / Q000378:metabolism; D012689:Sequence Homology, Nucleic Acid; D013379:Substrate Specificity; D000516:alpha-Amylases / Q000235:genetics* / Q000378:metabolism
D001426:Bacterial Proteins; D006026:Glycoside Hydrolases; D000516:alpha-Amylases; C100181:pullulanase