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J Struct Biol. 2016 Dec;196(3):309-318. doi: 10.1016/j.jsb.2016.07.017. Epub 2016 Jul 27.

Small-angle X-ray scattering of calpain-5 reveals a highly open conformation among calpains.

Author information

1
Department of Biochemistry, University of Iowa, Iowa City, IA, USA; Protein Crystallography Facility, University of Iowa, Iowa City, IA, USA.
2
Department of Pediatrics, University of Iowa, Iowa City, IA, USA; Omics Lab, University of Iowa, Iowa City, IA, USA.
3
Omics Lab, University of Iowa, Iowa City, IA, USA; Department of Ophthalmology and Visual Sciences, University of Iowa, Iowa City, IA, USA; Medical Scientist Training Program, University of Iowa, Iowa City, IA, USA.
4
Protein Crystallography Facility, University of Iowa, Iowa City, IA, USA.
5
Barbara and Donald Jonas Laboratory of Stem Cells and Regenerative Medicine and Bernard & Shirlee Brown Glaucoma Laboratory, Edward S. Harkness Eye Institute, Columbia University, New York, NY, USA; Department of Pathology & Cell Biology, College of Physicians & Surgeons, Columbia University, New York, NY, USA.
6
Omics Lab, University of Iowa, Iowa City, IA, USA; Department of Ophthalmology and Visual Sciences, University of Iowa, Iowa City, IA, USA. Electronic address: mahajanlab@gmail.com.

Abstract

Calpain-5 is a calcium-activated protease expressed in the retina. Mutations in calpain-5 cause autosomal dominant neovascular inflammatory vitreoretinopathy (ADNIV, OMIM#193235). The structure of calpain-5 has not been determined, thus hindering the investigation of its proteolytic targets and pathological role in ADNIV. Herein, we report models of the proteolytic core of calpain-5 (mini-calpain-5) containing two globular domains (termed DIIa-IIb) connected by a short, flexible linker, consistent with small-angle X-ray scattering (SAXS) data. Structural modeling in the absence of calcium suggests that mini-calpain-5 adopts a more open conformation when compared to previously determined structures of other calpain cores. This open conformation, achieved by a rotation of DIIa and DIIb with respect to each other, prevents formation of the active site and constrains the enzyme in an inactivated form. The relative domain rotation of 60-100° we found for mini-calpain-5 (a non-classical calpain) is significantly greater than the largest rotation previously observed for a classical calpain (i.e., 55.0° for mini-calpain-9). Together with our prediction that, in the full-length form, a long loop in DIIb (loop C1), a few residues downstream of the inter-domain linker, likely interacts with the shorter, acidic, inactivating loop on domain-III (DIII), these structural insights illuminate the complexity of calpain regulation. Moreover, our studies argue that pursuing higher resolution structural studies are necessary to understand the complex activity regulation prevalent in the calpain family and for the design of specific calpain inhibitors.

KEYWORDS:

ADNIV; CAPN5; Protease; SAXS; Uveitis

PMID:
27474374
PMCID:
PMC5118095
DOI:
10.1016/j.jsb.2016.07.017
[Indexed for MEDLINE]
Free PMC Article

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