The specificity and elastinolytic activities of bovine cathepsins S and H

Xin, Xue-Qin; Gunesekera, Bhadra; Mason, Robert W.

doi:10.1016/0003-9861(92)90283-3

Cited by 58 publications

(39 citation statements)

References 20 publications

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“…The other residues showing a minor preference at this position were Phe, Ile, and Val (and Lys for the P 1 ϭ Arg sublibrary). Single substrate experiments with bovine cathepsin H showed a preference for aliphatic hydrophobic residues over aromatic residues (67). A broad specificity was observed for the S 3 subsite.…”

Section: Resultsmentioning

confidence: 90%

High Throughput Substrate Specificity Profiling of Serine and Cysteine Proteases Using Solution-phase Fluorogenic Peptide Microarrays

Gosalia

Salisbury

Ellman

et al. 2005

Molecular & Cellular Proteomics

154

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Proteases regulate numerous biological processes with a degree of specificity often dictated by the amino acid sequence of the substrate cleavage site. To map protease/substrate interactions, a 722-member library of fluorogenic protease substrates of the general format AcAla-X-X-(Arg/Lys)-coumarin was synthesized (X ‫؍‬ all natural amino acids except cysteine) and microarrayed with fluorescent calibration standards in glycerol nanodroplets on glass slides. Specificities of 13 serine proteases (activated protein C, plasma kallikrein, factor VIIa, factor IXa␤, factor XIa and factor ␣ XIIa, activated complement C1s, C1r, and D, tryptase, trypsin, subtilisin Carlsberg, and cathepsin G) and 11 papain-like cysteine proteases (cathepsin B, H, K, L, S, and V, rhodesain, papain, chymopapain, ficin, and stem bromelain) were obtained from 103,968 separate microarray fluorogenic reactions (722 substrates ؋ 24 different proteases ؋ 6 replicates). This is the first comprehensive study to report the substrate specificity of rhodesain, a papain-like cysteine protease expressed by Trypanasoma brucei rhodesiense, a parasitic protozoa responsible for causing sleeping sickness. Rhodesain displayed a strong P 2 preference for Leu, Val, Phe, and Tyr in both the P 1 ‫؍‬ Lys and Arg libraries. Solution-phase microarrays facilitate protease/substrate specificity profiling in a rapid manner with minimal peptide library or enzyme usage. Molecular & Cellular Proteomics 4:626 -636, 2005.Because of their critical roles in biological pathways like hormone activation, proteasomal degradation, and apoptosis, proteases are essential for cellular function and viability. Proteases regulate hormonal activation, cellular homeostasis, apoptosis, and coagulation and play an important role in the pathogenicity and progression of many diseases (1). Proteases comprise one of the largest protein families in organisms from Escherichia coli to humans (2-4). Improved understanding of proteases will provide insight into biological systems and will likely provide a number of important new therapeutic targets (1).To properly function, proteases must preferentially cleave their target substrates in the presence of other proteins. While many factors impact protease substrate selection, one of the key aspects is the complementary nature of the enzymeactive site with the residues surrounding the cleaved bond in the substrate. As such, determination of the residues that comprise the preferred cleavage site of a protease provides critical information regarding substrate selection. Furthermore, determination of substrate specificity also provides a framework for the design of potent and selective inhibitors.Here we exploit solution-phase substrate nanodroplet microarrays (5), in which fluorogenic substrates suspended in glycerol droplets are treated with aerosolized aqueous enzyme solutions, to provide protease substrate specificity profiles (6). These arrays allow high throughput characterization of the preferred residues on the P side (7) of the substrate in a highl...

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Section: Resultsmentioning

confidence: 90%

High Throughput Substrate Specificity Profiling of Serine and Cysteine Proteases Using Solution-phase Fluorogenic Peptide Microarrays

Gosalia

Salisbury

Ellman

et al. 2005

Molecular & Cellular Proteomics

154

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“…[17][18][19] The time frame of the changes in cathepsin S and K expression and the local- The relative amounts of elastin degradation are presented as the absorbance determined. Elastolytic activity of cathepsin S (100 nmol) was used as a positive control.…”

Section: Discussionmentioning

confidence: 99%

“…16 Despite their lysosomal origin and optimal acidic pH, some of cathepsins including cathespin S and K can be secreted and retain a large portion of their proteolytic activity at neutral pH. [17][18][19] Among the members of the cathepsin family, cathepsin S and K express potent elastolytic as well as collagenolytic activities. 19 -21 Although it has been demonstrated that vascular SMCs have the ability to express these cathespins, 6,22 cathepsins have received much less consideration in the involvement in the neointima formation.…”

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confidence: 99%

Increased Expression of Elastolytic Cysteine Proteases, Cathepsins S and K, in the Neointima of Balloon-Injured Rat Carotid Arteries

Cheng

Kuzuya

Sasaki

et al. 2004

The American Journal of Pathology

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The matrix-degrading activity of several proteases are involved in the accelerated breakdown of extracellular matrix associated with vascular remodeling during the development of atherosclerosis and vascular injury-induced neointimal formation. Previous studies have shown that the potent elastolytic cysteine proteases, cathepsins S and K, are overexpressed in atherosclerotic lesions in human and animal models. However, the role of these cathepsins in vascular remodeling remains unclear. In the present study, the expressions of cathepsin S and K and their inhibitor cystatin C were examined during arterial remodeling using a rat carotid artery balloon-injury model. The increase in both cathepsin S and K mRNA levels was observed from day 1 and day 3 through day 14 following the induction of balloon injury, respectively. Western blotting analysis revealed that both cathepsin S and K protein levels also increased in the carotid arteries during neointima formation, coinciding with an increase elastolytic activity assayed using Elastin-Congo red, whereas, no significant change in the expressions of cystatin C mRNA and protein was observed during follow-up periods after injury. Immunohistochemistry, Western blot, and in situ hybridization showed that the increase of cathepins S and K and the decrease of cystatin C occurred preferentially in the developing neointima. These findings suggest that cathepsin S and K may participate in the pathological arterial remodeling associated with restenosis.

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“…Cathepsins are thought to be involved in the pathological degradation of extracellular matrix proteins like collagen [19], proteoglycans [20] and elastin [21]. As such, specific inhibition of these proteases represents a potentially important therapeutic strategy.…”

Section: Introductionmentioning

confidence: 99%

Peptidyl vinyl sulphones: a new class of potent and selective cysteine protease inhibitors: S2P2 specificity of human cathepsin O2 in comparison with cathepsins S and L

Brὂmme

Klaus

Okamoto

et al. 1996

Biochemical Journal

136

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Peptidyl vinyl sulphones are a novel class of extremely potent and specific cysteine protease inhibitors. They are highly active against the therapeutically important cathepsins O2, S and L. The highest k inact \K i values exceed 10( M −" :s −" for cathepsin S and 10& M −" :s −" for cathepsins O2 and L. To study the primary specificity site of the novel human cathepsin O2 and the effectiveness of this novel class of inhibitors, a series of peptidyl vinyl sulphones with variations in the P # residue was synthesized. Leucine in the P # position was proven to be the most effective residue for cathepsin O2 and also for cathepsins S and L.

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The specificity and elastinolytic activities of bovine cathepsins S and H

Cited by 58 publications

References 20 publications

High Throughput Substrate Specificity Profiling of Serine and Cysteine Proteases Using Solution-phase Fluorogenic Peptide Microarrays

High Throughput Substrate Specificity Profiling of Serine and Cysteine Proteases Using Solution-phase Fluorogenic Peptide Microarrays

Increased Expression of Elastolytic Cysteine Proteases, Cathepsins S and K, in the Neointima of Balloon-Injured Rat Carotid Arteries

Peptidyl vinyl sulphones: a new class of potent and selective cysteine protease inhibitors: S2P2 specificity of human cathepsin O2 in comparison with cathepsins S and L

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