Differences in transition state stabilization between thermolysin (EC 3.4.24.27) and neprilysin (EC 3.4.24.11).
Important homologies in the topology of the catalytic site and the mechanism of action of thermolysin and neprilysin have been evidenced by site-directed mutagenesis. The determination of differences in transition state stabilization between these peptidases could facilitate the design of specific inhibitors. Thus, two residues of thermolysin which could be directly (Tyr157) or indirectly (Asp226) involved in the stabilization of the transition state and their putative counterparts in neprilysin (Tyr625 and Asp709) have been mutated. The results show that Tyr157 is important for thermolysin activity while Tyr625 has no functional role in neprilysin. Conversely, the mutation of Asp226 induced a slight perturbation of thermolysin activity while Asp709 in neprilysin seems crucial in neprilysin catalysis. Taken together these data seem to indicate differences in the transition state mode of stabilization in the two peptidases.
Marie-Claire
C
C
Département de Pharmacochimie Moléculaire et Structurale, U266 INSERM, URA D1500 CNRS, UFR des Sciences Pharmaceutiques et Biologiques, Faculté de Pharmacie, Paris, France.
Ruffet
E
E
Tiraboschi
G
G
Fournie-Zaluski
M C
MC
D000595:Amino Acid Sequence; D001224:Aspartic Acid; D001412:Bacillus subtilis / Q000201:enzymology; D020134:Catalytic Domain; D004795:Enzyme Stability; D006020:Glycopeptides / Q000494:pharmacology; D007700:Kinetics; D008958:Models, Molecular; D016297:Mutagenesis, Site-Directed; D015260:Neprilysin / Q000737:chemistry* / Q000378:metabolism*; D011480:Protease Inhibitors / Q000494:pharmacology; D011487:Protein Conformation; D011994:Recombinant Proteins / Q000737:chemistry / Q000378:metabolism; D013820:Thermolysin / Q000737:chemistry* / Q000378:metabolism*; D015244:Thiorphan / Q000494:pharmacology; D014443:Tyrosine
D006020:Glycopeptides; D011480:Protease Inhibitors; D011994:Recombinant Proteins; D001224:Aspartic Acid; D014443:Tyrosine; D015244:Thiorphan; D015260:Neprilysin; D013820:Thermolysin; C008890:phosphoramidon