Identification of serpin determinants of specificity and selectivity for furin inhibition through studies of α1PDX (α1-protease inhibitor Portland)-serpin B8 and furin active-site loop chimeras.
journal contributionposted on 2014-07-04, 00:00 authored by Gonzalo Izaguirre, Lixin Qi, Mary Lima, Steven T. Olson
α1-Protease inhibitor Portland (α1PDX) is an engineered serpin family inhibitor of the proprotein convertase (PC), furin, that exhibits high specificity but limited selectivity for inhibiting furin over other PC family proteases. Here, we characterize serpin B8, a natural inhibitor of furin, together with α1PDX-serpin B8 and furin-PC chimeras to identify determinants of serpin specificity and selectivity for furin inhibition. Replacing reactive center loop (RCL) sequences of α1PDX with those of serpin B8 demonstrated that both the P4-P1 RXXR recognition sequence as well as the P1'-P5' sequence are critical determinants of serpin specificity for furin. Alignments of PC catalytic domains revealed four variable active-site loops whose role in furin reactivity with serpin B8 was tested by engineering furin-PC loop chimeras. The furin(298-300) loop but not the other loops differentially affected furin reactivity with serpin B8 and α1PDX in a manner that depended on the serpin RCL-primed sequence. Modeling of the serpin B8-furin Michaelis complex identified serpin exosites in strand 3C close to the 298-300 loop whose substitution in α1PDX differentially affected furin reactivity depending on the furin loop and serpin RCL-primed sequences. These studies demonstrate that RCL-primed residues, strand 3C exosites, and the furin(298-300) loop are critical determinants of serpin reactivity with furin, which may be exploited in the design of specific and selective α1PDX inhibitors of PCs.
This work was supported by NIH grant R37-HL39888
Publisher StatementThis research was originally published in Journal of Biological Chemistry . Gonzalo Izaguirre, Lixin Qi, Mary Lima, and Steven T. Olson. Identification of serpin determinants of specificity and selectivity for furin inhibition through studies of α1PDX (α1-protease inhibitor Portland)-serpin B8 and furin active-site loop chimeras. Journal of Biological Chemistry . 2013. 288:21802-14. © the American Society for Biochemistry and Molecular Biology
PublisherAmerican Society for Biochemistry and Molecular Biology