Multiple domains of endopin 2A for serpin cross-class inhibition of papain.
The serpin endopin 2A inhibits the cysteine protease papain in cross-class inhibition. This study demonstrates the novel finding that both the non-RSL NH(2)-domain and the RSL domain with P1-P1' residues participate in endopin 2A inhibition. Production of a chimeric mutant of endopin 2A with replacement of its NH(2)-domain with that of endopin 1 resulted in less effective inhibition of papain, indicated by its lower k(ass) association rate constant compared to wild-type endopin 2A. This chimeric mutant formed complexes with papain, but at lower levels compared to that with wild-type endopin 2A. Papain degradation of a portion of the chimeric mutant suggested a role for the NH(2)-domain in regulating relative amounts of endopin 2A that enter the substrate pathway compared to the serpin inhibitory pathway. Furthermore, site-directed mutagenesis demonstrated that the RSL domain with intact P1-P1' residues was necessary for inhibition. These findings indicate that the NH(2)-domain and the RSL region both participate in endopin 2A inhibition of papain.
Hwang
Shin-Rong
SR
Skaggs School of Pharmacy and Pharmaceutical Sciences, University of California, San Diego, La Jolla, CA 92093-0744, USA.
Hook
Vivian Y H
VY
D000818:Animals; D001483:Base Sequence; D001665:Binding Sites / Q000235:genetics; D002417:Cattle; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D010206:Papain / Q000037:antagonists & inhibitors* / Q000737:chemistry; D017434:Protein Structure, Tertiary / Q000235:genetics; D015843:Serpins / Q000737:chemistry* / Q000235:genetics / Q000502:physiology*
D015843:Serpins; C412321:endopin 2; D010206:Papain