Structural adaptation to selective pressure for altered ligand specificity in the Pseudomonas aeruginosa amide receptor, amiC.
The AmiC protein in Pseudomonas aeruginosa is the negative regulator and ligand receptor for an amide-inducible aliphatic amidase operon. In the wild-type PAC1 strain, amidase expression is induced by acetamide or lactamide, but not by butyramide. A mutant strain of P. aeruginosa, PAC181, was selected for its sensitivity to induction by butyramide. The molecular basis for the butyramide inducible phenotype of P.aeruginosa PAC181 has now been determined, and results from a Thr-->Asn mutation at position 106 in PAC181-AmiC. In the wild-type PAC1-AmiC protein this residue forms part of the side wall of the amide-binding pocket but does not interact with the acetamide ligand directly. In the crystal structure of PAC181-AmiC complexed with butyramide, the Thr-->Asn mutation increases the size of the ligand binding site such that the mutant protein is able to close into its 'on' configuration even in the presence of butyramide. Although the mutation allows butyramide to be recognized as an inducer of amidase expression, the mutation is structurally sub-optimal, and produces a significant decrease in the stability of the mutant protein.
O'Hara
B P
BP
Department of Biochemistry and Molecular Biology and Joint UCL/LICR X-Ray Crystallography Laboratory, University College London, Gower Street, London WC1E 6BT, UK.
Wilson
S A
SA
Lee
A W
AW
Roe
S M
SM
Siligardi
G
G
Drew
R E
RE
Pearl
L H
LH
D000220:Adaptation, Biological / Q000235:genetics*; D000577:Amides / Q000378:metabolism / Q000494:pharmacology; D000581:Amidohydrolases / Q000235:genetics / Q000378:metabolism; D019943:Amino Acid Substitution; D001426:Bacterial Proteins / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism; D001665:Binding Sites; D002942:Circular Dichroism; D018360:Crystallography, X-Ray; D005809:Genes, Regulator; D006358:Hot Temperature; D006860:Hydrogen Bonding; D008024:Ligands; D008958:Models, Molecular; D020125:Mutation, Missense; D033902:Periplasmic Binding Proteins; D010641:Phenotype; D011485:Protein Binding; D011489:Protein Denaturation; D015202:Protein Engineering; D017434:Protein Structure, Tertiary; D011550:Pseudomonas aeruginosa / Q000737:chemistry* / Q000235:genetics*; D012097:Repressor Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D012641:Selection, Genetic
D000577:Amides; D001426:Bacterial Proteins; D008024:Ligands; D033902:Periplasmic Binding Proteins; D012097:Repressor Proteins; C070135:AmiC protein, Pseudomonas aeruginosa; C006866:butyramide; D000581:Amidohydrolases; C019665:amidase