Engineering of halophilic enzymes: two acidic amino acid residues at the carboxy-terminal region confer halophilic characteristics to Halomonas and Pseudomonas nucleoside diphosphate kinases.
Nucleoside diphosphate kinase from Halomonas sp. 593 (HaNDK) exhibits halophilic characteristics. Residues 134 and 135 in the carboxy-terminal region of HaNDK are Glu-Glu, while those of its homologous counterpart of non-halophilic Pseudomonas NDK (PaNDK) are Ala-Ala. The double mutation, E134A-E135A, in HaNDK results in the loss of the halophilic characteristics, and, conversely, the double mutation of A134E-A135E in PaNDK confers halophilic characters to this enzyme, indicating that the charged state of these two residues that are located in the C-terminal region plays a critical role in determining halophilic characteristics. The importance of these two residues versus the net negative charges will be discussed in relation to the halophilicity of NDK.
Tokunaga
Hiroko
H
Applied and Molecular Microbiology, Faculty of Agriculture, Kagoshima University, Kagoshima 890-0065, Japan.
Arakawa
Tsutomu
T
Tokunaga
Masao
M
D000595:Amino Acid Sequence; D024342:Amino Acids, Acidic / Q000235:genetics / Q000378:metabolism*; D004305:Dose-Response Relationship, Drug; D004795:Enzyme Stability; D004926:Escherichia coli / Q000235:genetics; D020632:Halomonas / Q000201:enzymology*; D006358:Hot Temperature; D007526:Isoelectric Point; D008969:Molecular Sequence Data; D008970:Molecular Weight; D009154:Mutation; D009701:Nucleoside-Diphosphate Kinase / Q000737:chemistry* / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism; D011489:Protein Denaturation; D015202:Protein Engineering; D020673:Protein Renaturation; D017433:Protein Structure, Secondary; D011549:Pseudomonas / Q000201:enzymology*; D012965:Sodium Chloride / Q000494:pharmacology; D012995:Solubility; D013997:Time Factors
D024342:Amino Acids, Acidic; D012965:Sodium Chloride; D009701:Nucleoside-Diphosphate Kinase