Crystal structure of human osteoclast cathepsin K complex with E-64

Zhao, Baoguang; Janson, Cheryl A.; Amegadzie, Bernard Y.; D'Alessio, Karla; Griffin, Charles W.; Hanning, Charles R.; Jones, Christopher S.; Kurdyla, Jeffrey T.; McQueney, Michael S.; Qiu, Xiayang; Smith, Ward W.; Abdel-Meguid, Sherin S.

doi:10.1038/nsb0297-109

Cited by 118 publications

(92 citation statements)

References 20 publications

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“…In papain family cysteine proteases, the binding of E-64 to the catalytic cysteine, which is located at the bottom of the substrate binding cleft, has been reported to induce only a small structural changes (55)(56)(57)(58). This is considered to be the case with Der p 1, and E-64 covers only a small area of the Der p 1 in the model of Der p 1/E-64 complex (Fig.…”

Section: E-64-treated Rder P 1 Retained Its Global Structurementioning

confidence: 99%

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Crucial Commitment of Proteolytic Activity of a Purified Recombinant Major House Dust Mite Allergen Der p1 to Sensitization toward IgE and IgG Responses

Kikuchi

Takai²,

Kuhara³

et al. 2006

The Journal of Immunology

102

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The major proteolytic allergen derived from the house dust mite Dermatophagoides pteronyssinus, Der p1, is one of the most clinically relevant allergens worldwide. In the present study, we evaluate the contribution of the proteolytic activity and structure of a highly purified rDer p 1 to immune responses. Mice were i.p. immunized with three forms of rDer p 1 adsorbed to Alum: one enzymatically active, one treated with an irreversible cysteine protease-specific inhibitor, E-64, and one heat denatured. Immunization with E-64-treated or heat-denatured rDer p 1 elicited much less production of serum total IgE and not only rDer p 1-specific IgE but also IgGs compared with immunization with active rDer p 1. Assays for Ab-binding and its inhibition and structural analyses indicated that E-64-treated rDer p 1 retained its global structure and conformational B cell epitopes. A proliferative response and production of IL-5 by spleen cells restimulated with rDer p 1 were observed on immunization with the active rDer p 1 but not E-64-treated rDer p 1. The cells from mice immunized with heat-denatured rDer p 1 exhibited the highest levels of proliferation and production of IL-5 and IFN-γ. The results indicate that the proteolytic activity of the highly purified rDer p 1 crucially commits to the sensitization process, including both IgE and IgG responses. Additionally, we demonstrated immunogenic differences by functional or structural manipulations of the rDer p 1. The findings have implications for sensitization to this relevant allergen in humans and for the design of modified allergen-vaccines for future allergen-specific immunotherapy.

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Section: E-64-treated Rder P 1 Retained Its Global Structurementioning

confidence: 99%

“…A model of the Der p 1/E-64 complex was made by superimposing the Protein Data Bank entries 1XKG (a mutant of pro-Der p 1) (20) and 1ATK (cathepsin K/E-64 complex) (55) and removing unwanted portions. The figure was made by PyMOL (DeLano Scientific).…”

Section: Molecular Modelingmentioning

confidence: 99%

Crucial Commitment of Proteolytic Activity of a Purified Recombinant Major House Dust Mite Allergen Der p1 to Sensitization toward IgE and IgG Responses

Kikuchi

Takai²,

Kuhara³

et al. 2006

The Journal of Immunology

102

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“…X-ray diffraction data were measured from a single crystal by using a Siemens two-dimensional, position-sensitive detector on a Siemens rotating anode generator operating at 5 kW. The structure was determined by rigid body refinement by using X-PLOR, and the starting model consisted of the protein atoms from the crystal structure of cathepsin K in complex with the cysteine protease inhibitor E64 (16). Fourier maps with coefficients ͉F o -F c ͉ and ͉2F o -F c ͉ were used to fit the atomic model 1 2, with cell constants of a ϭ 57.6 Å and c ϭ 131.2 Å.…”

Section: Methodsmentioning

confidence: 99%

“…Protein was prepared as described previously (16). Crystals of mature, activated cathepsin K complexed with inhibitor 4 grew to a size of Ϸ0.2 mm 3 in about 6 days at 20°C.…”

Section: Methodsmentioning

confidence: 99%

Design of potent and selective human cathepsin K inhibitors that span the active site

Thompson

Halbert

Bossard

et al. 1997

Proc. Natl. Acad. Sci. U.S.A.

Self Cite

127

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Potent and selective active-site-spanning inhibitors have been designed for cathepsin K, a cysteine protease unique to osteoclasts. They act by mechanisms that involve tight binding intermediates, potentially on a hydrolytic pathway. X-ray crystallographic, MS, NMR spectroscopic, and kinetic studies of the mechanisms of inhibition indicate that different intermediates or transition states are being represented that are dependent on the conditions of measurement and the specific groups f lanking the carbonyl in the inhibitor. The species observed crystallographically are most consistent with tetrahedral intermediates that may be close approximations of those that occur during substrate hydrolysis. Initial kinetic studies suggest the possibility of irreversible and reversible active-site modification. Representative inhibitors have demonstrated antiresorptive activity both in vitro and in vivo and therefore are promising leads for therapeutic agents for the treatment of osteoporosis. Expansion of these inhibitor concepts can be envisioned for the many other cysteine proteases implicated for therapeutic intervention.

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“…The models of complexes of testican Tg-1 domain with papain and cathepsins K and L were calculated with the homology modelling program Modeler (Š ali and Blundell, 1993) using the crystal structures of papain (Kamphuis et al, 1984), cathepsin K (Zhao et al, 1997) and p41 fragment complexed with cathepsin L (Gunč ar et al, 1999) as templates (PDB codes 9PAP, 1ATK and 1ICF, respectively). Prior to modelling, the template structures were cleaned of water molecules and non-native atoms (oxygen atoms on oxidised Cys 25 in papain and E-64 in the cathepsin K structure).…”

Section: Homology Modellingmentioning

confidence: 99%

Dual concentration-dependent activity of thyroglobulin type-1 domain of testican: specific inhibitor and substrate of cathepsin L

Meh

Pavšič

Türk

et al. 2005

Biological Chemistry

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The thyroglobulin type-1 (Tg-1) domain is a protein module that occurs in a variety of secreted and membrane proteins and is recognised as a potent inhibitor of cysteine peptidases. We present here some properties of the Tg-1 domain of human testican, a modularly organised proteoglycan secreted mainly by brain cells, the exact in vivo function of which is not yet clear. The domain was prepared as a recombinant protein in a Pichia pastoris expression system and its activity was demonstrated by specific and selective inhibition of cathepsin L (K i s0.14 nM). Interaction at high enzyme and inhibitor concentrations resulted in degradation of the domain by cathepsin L, which was not observed under conditions used for the determination of kinetic parameters. No inhibitory activity could be detected for cathepsin K, but it exhibited a very similar degradation pattern. Homology modelling provided a good explanation for the different behaviour observed with the two enzymes. Firstly, the steric fit between the interfaces of testican domain and cathepsin L is stabilised by numerous favourable forces, while no such interactions are evident in the complex with cathepsin K, and repulsive interactions even prevent access of the domain to the active site of papain. Secondly, the prolonged first loop of the domain occupies a position near the catalytic cysteine residue in a more substrate-like manner, enabling cleavage of the Gly 22 -Ala 23 bond.

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Crystal structure of human osteoclast cathepsin K complex with E-64

Cited by 118 publications

References 20 publications

Crucial Commitment of Proteolytic Activity of a Purified Recombinant Major House Dust Mite Allergen Der p1 to Sensitization toward IgE and IgG Responses

Crucial Commitment of Proteolytic Activity of a Purified Recombinant Major House Dust Mite Allergen Der p1 to Sensitization toward IgE and IgG Responses

Design of potent and selective human cathepsin K inhibitors that span the active site

Dual concentration-dependent activity of thyroglobulin type-1 domain of testican: specific inhibitor and substrate of cathepsin L

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