Structural basis for the inactivation of the P54 mutant of beta-lactamase from Staphylococcus aureus PC1.
The crystal structure of a mutant protein of a class A beta-lactamase from Staphylococcus aureus PC1, in which Asp179 is replaced by an asparagine (P54), has been determined and refined at 2.3-A resolution (1 A = 0.1 nm). The resulting crystallographic R factor [formula: see text] are the observed and calculated structure factor amplitudes) is 0.181 for 12289 reflections with I greater than or equal to sigma (I) within the 6.0-2.3-A resolution range. The mutated residue is located at the C-terminus of an extensive loop (the omega-loop), remote from the active site, and results in a drastically reduced activity. Examination of the native and P54 structures reveals that the overall fold is similar, except that there is substantial disorder of the omega-loop of P54. This is a consequence of the elimination of a salt bridge between Asp179 and Arg164 that links the two ends of the omega-loop in native beta-lactamase. It is associated with a difference in side-chain conformation between Asn179 in P54 and Asp179 in the native structure. An alternate interaction occurs in P54 between Asn179 and Ala69, adjacent to the catalytic Ser70. This disorder affects catalysis since some of the disordered residues, in particular Glu166, form part of the active site.(ABSTRACT TRUNCATED AT 250 WORDS)
Herzberg
O
O
Center for Advanced Research in Biotechnology, Maryland Biotechnology Institute, University of Maryland, Rockville 20850.
Kapadia
G
G
Blanco
B
B
Smith
T S
TS
Coulson
A
A
D000645:Ammonium Sulfate / Q000494:pharmacology; D001665:Binding Sites; D003461:Crystallography; D004252:DNA Mutational Analysis; D006863:Hydrogen-Ion Concentration; D007700:Kinetics; D008958:Models, Molecular; D010400:Penicillin G / Q000378:metabolism; D011487:Protein Conformation; D013211:Staphylococcus aureus / Q000201:enzymology*; D013329:Structure-Activity Relationship; D014961:X-Ray Diffraction; D001618:beta-Lactamases / Q000737:chemistry* / Q000235:genetics
D001618:beta-Lactamases; D010400:Penicillin G; D000645:Ammonium Sulfate