title: Raw starch-degrading α-amylase from Bacillus aquimaris MKSC 6.2: isolation and expression of the gene, bioinformatics and biochemical characterization of the recombinant enzyme.
abstract: OBJECTIVE
  The aims were to isolate a raw starch-degrading α-amylase gene baqA from Bacillus aquimaris MKSC 6.2, and to characterize the gene product through in silico study and its expression in Escherichia coli.
  
  
  RESULTS
  A 1539 complete open reading frame of a starch-degrading α-amylase gene baqA from B. aquimaris MKSC 6·2 has been determined by employing PCR and inverse PCR techniques. Bioinformatics analysis revealed that B. aquimaris MKSC 6.2 α-amylase (BaqA) has no starch-binding domain, and together with a few putative α-amylases from bacilli may establish a novel GH13 subfamily most closely related to GH13_1. Two consecutive tryptophans (Trp201 and Trp202, BaqA numbering) were identified as a sequence fingerprint of this novel GH13 subfamily. Escherichia coli cells produced the recombinant BaqA protein as inclusion bodies. The refolded recombinant BaqA protein degraded raw cassava and corn starches, but exhibited no activity with soluble starch.
  
  
  CONCLUSIONS
  A novel raw starch-degrading B. aquimaris MKSC 6.2 α-amylase BaqA is proposed to be a member of new GH13 subfamily.
  
  
  CONCLUSIONS
  This study has contributed to the overall knowledge and understanding of amylolytic enzymes that are able to bind and digest raw starch directly.
authors.affiliation: Biochemistry Research Division, Faculty of Mathematics and Natural Sciences, Institut Teknologi Bandung, Bandung, Indonesia.
authors.forename: F
authors.initials: F
authors.lastname: Puspasari
authors.forename: O K
authors.initials: OK
authors.lastname: Radjasa
authors.forename: A S
authors.initials: AS
authors.lastname: Noer
authors.forename: Z
authors.initials: Z
authors.lastname: Nurachman
authors.forename: Y M
authors.initials: YM
authors.lastname: Syah
authors.forename: M
authors.initials: M
authors.lastname: van der Maarel
authors.forename: L
authors.initials: L
authors.lastname: Dijkhuizen
authors.forename: S
authors.initials: S
authors.lastname: Janeček
authors.forename: D
authors.initials: D
authors.lastname: Natalia
mesh_terms: D000595:Amino Acid Sequence; D001407:Bacillus / Q000201:enzymology* / Q000235:genetics; D001483:Base Sequence; D019295:Computational Biology; D004926:Escherichia coli / Q000235:genetics / Q000378:metabolism; D005798:Genes, Bacterial; D008969:Molecular Sequence Data; D016366:Open Reading Frames; D017510:Protein Folding; D054730:Protein Interaction Domains and Motifs; D017434:Protein Structure, Tertiary; D011994:Recombinant Proteins / Q000235:genetics / Q000378:metabolism; D013213:Starch / Q000378:metabolism*; D000516:alpha-Amylases / Q000235:genetics* / Q000378:metabolism
chemical_list: D011994:Recombinant Proteins; D013213:Starch; D000516:alpha-Amylases