title: Role of two amino acid residues' insertion on thermal stability of thermophilic α-amylase AMY121 from a deep sea bacterium Bacillus sp. SCSIO 15121.
abstract: α-Amylases from Bacillus licheniformis (BLA) and Bacillus amyloliquefaciens (BAA) are both important industrial enzymes with high similarity in structure but significant differences in thermostability. The mechanisms underlying this discrepancy are still poorly understood. Here, we investigated the role of two amino acids' insertion on the thermostability of these two group amylases. A newly obtained thermophilic amylase AMY121 was found much closer to BLA in both primary structure and enzymological properties. Two amino acids' insertion widespread among BAA group α-amylases was identified as one of the key factors leading to the thermostability differences, since thermostability of insertion mutants (AMY121-EG and AMY121-AA) from AMY121 significantly decreased, while that of deletion mutant from BAA increased. Moreover, we proposed that conformational disturbance caused by insertion mutation might weaken the calcium-binding affinity and consequently decrease the enzyme thermostability.
authors.affiliation: CAS Key Laboratory of Tropical Marine Bio-resources and Ecology, Guangdong Key Laboratory of Marine Materia Medica, RNAM Center for Marine Microbiology, South China Sea Institute of Oceanology, Chinese Academy of Sciences, Guangzhou, 510301, People's Republic of China.
authors.forename: Lizhen
authors.initials: L
authors.lastname: Li
authors.forename: Jian
authors.initials: J
authors.lastname: Yang
authors.forename: Jie
authors.initials: J
authors.lastname: Li
authors.forename: Lijuan
authors.initials: L
authors.lastname: Long
authors.forename: Yunzhu
authors.initials: Y
authors.lastname: Xiao
authors.forename: Xinpeng
authors.initials: X
authors.lastname: Tian
authors.forename: Fazuo
authors.initials: F
authors.lastname: Wang
authors.forename: Si
authors.initials: S
authors.lastname: Zhang
mesh_terms: D000595:Amino Acid Sequence; D000596:Amino Acids / Q000737:chemistry; D001407:Bacillus / Q000201:enzymology*; D001665:Binding Sites; D002118:Calcium / Q000737:chemistry; D003001:Cloning, Molecular; D004795:Enzyme Stability / Q000235:genetics; D004926:Escherichia coli / Q000378:metabolism; D017353:Gene Deletion; D006863:Hydrogen-Ion Concentration; D007477:Ions; D008670:Metals / Q000737:chemistry; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D009154:Mutation; D010802:Phylogeny; D011487:Protein Conformation; D011994:Recombinant Proteins / Q000737:chemistry; D013696:Temperature; D014871:Water Microbiology; D000516:alpha-Amylases / Q000737:chemistry*
chemical_list: D000596:Amino Acids; D007477:Ions; D008670:Metals; D011994:Recombinant Proteins; D000516:alpha-Amylases; D002118:Calcium