Active center differences between cathepsins L and B: The S<sub>1</sub> binding region

Kirschke, Heidrun; Wikström, P; Shaw, Elliott

doi:10.1016/0014-5793(88)80600-8

Cited by 64 publications

(36 citation statements)

References 25 publications

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“…2 ) and N-benzyloxycarbonyl-phenylalanyl-tyrosine(OBut)-diazomethane (CbzPhe-Tyr-(OBut)-CHN 2 ) were prepared by the general procedures (17,18) and had melting points in close agreement with the literature values (19). The biotinylated affinity label N-biotinyl-phenylalanyl-alanine-diazomethane (Bio-Phe-Ala-CHN 2 ) was prepared by the cited procedure (14).…”

Section: Synthesis Of Affinity Labelsmentioning

confidence: 75%

“…This activity could be completely inhibited by the general cysteine protease inhibitor iodoacetamide and, additionally, the activity was also inhibited by the broad spectrum affinity label for cysteine proteinases Cbz-Phe-Ala-CHN 2 (22), but not by the specific cathepsin L inhibitor Cbz-Phe-Tyr(OBut)-CHN 2 (19). The exhibited inhibitor susceptibility taken in consideration with the specificity of the Substrate (Cbz-ArgArg-N£UMec), suggests that the observed activity was indicative of a cathepsin B-like proteinäse and excludes the possibility that a ksdlikrein^like enzyme is responsible for the observed Substrate hydrolysis.…”

Section: Resultsmentioning

confidence: 99%

“…When samples of rheumatoid arthritic synovial fluid were pre-treated with Cbz-Phe-Tyr(OBut)-CHN 2 (0.1 mmol/1, for 30 min at 37 °C) which is just such an inactivator (19), no dimu> ution in the incorporation of the biotinylated probe was observed (compare lane 5 -pre-treated sample with and Biotin-Phe-Ala-CHN 2 Lane 6 synovial fluid pretreated with Cbz-Phe-Ala-CHN 2 and°B iotin-Phe-Ala-CHN 2 Lane 7 biotinylated molecular weight markers lane 3 -no pre-treatment). This further strengthens our case that the species of M T 42 000 and 29 000 are cathepsin B-like.…”

Section: Resultsmentioning

confidence: 99%

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The Detection, Quantification and Partial Characterisation of Cathepsin B-Like Activity in Human Pathological Synovial Fluids

Duffy¹,

Walker²,

Guthrie³

et al. 1994

CCLM

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Summary:In this study, the levels of the cysteine proteinase -cathepsin B were measured in diseased synovial fluids using a steady state fluorometric assay. Cathepsin B-like activity was shown to be present in all the samples analysed, with the rheumatoid arthritic synovial fluids possessing significantly higher concentrations (mean value ca. 416 mg/1) than the osteoarthritic fluids (mean value ca. 142.4 mg/1). In addition, upon treatment with pepsin, all of the rheumatoid arthritis samples were shown to possess additional cathepsin B-like activity, suggesting the presence of a reservoir of latent precursor molecules. By utilising a recently developed biotinylated affmity label for cathepsin B-like proteinases and sheep anti-(human cathepsin B) antibodies, used in combination with SDS-PAGE and Western blotting, the rheumatoid arthritic synovial cathepsin B was shown to exist in two forms with apparent molecular masses of M r 29 000 and 42 000. We propose that the former is a ftmctionally active proteinase, whereas the latter is a pepsin activatable proform which, when cleaved by this aspartyl proteinase, is converted into a catalytically competent species of M f 20000.

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Section: Synthesis Of Affinity Labelsmentioning

confidence: 75%

Section: Resultsmentioning

confidence: 99%

Section: Resultsmentioning

confidence: 99%

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The Detection, Quantification and Partial Characterisation of Cathepsin B-Like Activity in Human Pathological Synovial Fluids

Duffy¹,

Walker²,

Guthrie³

et al. 1994

CCLM

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“…Data are mean values from two experiments and showed a variation not greater than 3.5%. The preparation of the inhibitors are described in A, Crawford et al (1988); B, unpublished results; C, Shaw and Green (1981); D, Leary and Shaw (1977); E, Kirschke et al (1988);F, Green and Shaw (1981). Tyx, Tyr(O+Bu).…”

Section: Discussionmentioning

confidence: 99%

Characterisation of a cysteine protease from bloodstream forms of Trypanosoma congolense

Mbawa¹,

Gumm²,

Shaw

et al. 1992

European Journal of Biochemistry

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A cysteine protease (trypanopain-Tc) with cathepsin-L-like properties has been purified from Trypanosoma congolense. The enzyme has an apparent molecular mass of 31 -3 2 kDa by SDS/ PAGE and 66 kDa by gel chromatography. It has a PI 7.4 and a high affinity for concanavalin A, Trypanopain-Tc catalyses the limited proteolysis of a variety of protein substrates such as fibrinogen, serum albumin and trypanosome variant-surface glycoprotein. It has minimal or no activity against casein or elastin.A variety of peptidyl amidomethylcoumarins and peptidyl diazomethanes were used to test the specificity of trypanopain-Tc. The better substrates had Arg or Lys in P1 and hydrophobic amino acids in P2 and P3. The best substrate found for trypanopain-Tc was Z-Phe-Arg-NHMec (Z, benzyloxycarbonyl ; NHMec, 7-amido-4-methylcoumarin). The kinetic constants for the hydrolysis, of ZPhe-Arg-NHMec were k,,, = 17.4 SKI, K, = 4.4 pM, kcal/Km = 4.0 pM-' ' s-', which are very similar to those of cathepsin L with this substrate. The specific substrates for cathepsin B (Z-ArgArg-NHMec) and cathepsin H (Arg-NHMec) were not hydrolysed by trypanopain-Tc under the conditions tested. The pH optimum of trypanopain-Tc against Z-Phe-Arg-NHMec was pH 6.0 but it showed a broad peak of activity extending well into the alkaline region. The enzyme was activated by low-molecular-mass thiol compounds and inhibited by cystatin, ~-trans-epoxysucciny1-4-guanidinobutane (E-64) and a variety of peptidyl diazomethanes. The most effective diazomethane inhibitors (Z-Leu-Leu-Met-CHN2, Z-Leu-Met-CHN2 and Z-Leu-Lys-CHN2, were inhibitory at nanomolar concentrations and were trypanocidal in vituo after 24 -48 h incubation in 2 20 pM [inhibitor]. However, it is not clear whether the trypanocidal activity of these inhibitors is a consequence of the inhibition of trypanopains or of some other essential proteolytic activities within the parasites.Cathepsins L and B play important roles in intracellular protein turnover (for review see Ballard, 1987;Marzella and Glaumann, 1987). They are also implicated in the degradation of extracellular matrix in diseases such as muscular dystrophy, emphysema and arthritis, and in association with tumour metastasis (Rochefort ct al., 1987;Kominami et al., 1987; Burnett and Stockley, 1985;Etherington et al., 1988). Recently, interest has been shown in the role of cysteine proteases in the pathology of parasitic infections. These proteases can be released from the parasites (Blum, 1976;Keene, 1986; Lockwood et al., 1988; Garber and Lemchuk-Favel, 1989;North et al., 1989; Boutignon et al., 1990) andmay beinvolved in the pathology of the diseases and the survival of the parasites within the host (Bremmer et al

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“…2A). The cell-permeable cathepsin L inhibitor I (Z-PhePhe-FMK [benzyloxycarbonyl-Phe-Phe-fluoromethylketone]) (55,56) phenocopied the results observed with the more general inhibitors leupeptin and E-64, which may suggest that L1 is mostly processed by cathepsin L. In support of this notion, the generation of L1 cleavage products was only affected to a minor extent in the presence of the cell-permeable cathepsin B inhibitor CA-074Me that suppressed L1 I (57, 58) ( Fig. 2A).…”

Section: Hpv16 L1 Is Proteolytically Cleaved During Infectionmentioning

confidence: 99%

Kallikrein-8 Proteolytically Processes Human Papillomaviruses in the Extracellular Space To Facilitate Entry into Host Cells

Cerqueira

Ventayol

Vogeley

et al. 2015

J Virol

104

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The entry of human papillomaviruses into host cells is a complex process. It involves conformational changes at the cell surface, receptor switching, internalization by a novel endocytic mechanism, uncoating in endosomes, trafficking of a subviral complex to the Golgi complex, and nuclear entry during mitosis. Here, we addressed how the stabilizing contacts in the capsid of human papillomavirus 16 (HPV16) may be reversed to allow uncoating of the viral genome. Using biochemical and cell-biological analyses, we determined that the major capsid protein L1 underwent proteolytic cleavage during entry. In addition to a dispensable cathepsin-mediated proteolysis that occurred likely after removal of capsomers from the subviral complex in endosomes, at least two further proteolytic cleavages of L1 were observed, one of which was independent of the low-pH environment of endosomes. This cleavage occurred extracellularly. Further analysis showed that the responsible protease was the secreted trypsin-like serine protease kallikrein-8 (KLK8) involved in epidermal homeostasis and wound healing. Required for infection, the cleavage was facilitated by prior interaction of viral particles with heparan sulfate proteoglycans. KLK8-mediated cleavage was crucial for further conformational changes exposing an important epitope of the minor capsid protein L2. Occurring independently of cyclophilins and of furin that mediate L2 exposure, KLK8-mediated cleavage of L1 likely facilitated access to L2, located in the capsid lumen, and potentially uncoating. Since HPV6 and HPV18 also required KLK8 for entry, we propose that the KLK8-dependent entry step is conserved. IMPORTANCEOur analysis of the proteolytic processing of incoming HPV16, an etiological agent of cervical cancer, demonstrated that the capsid is cleaved extracellularly by a serine protease active during wound healing and that this cleavage was crucial for infection. The cleavage of L1 is one of at least four structural alterations that prime the virus extracellularly for receptor switching, internalization, and possibly uncoating. This step was also important for HPV6 and HPV18, which may suggest that it is conserved among the papillomaviruses. This study advances the understanding of how HPV16 initially infects cells, strengthens the notion that wounding facilitates infection of epidermal tissue, and may help the development of antiviral measures. Human papillomaviruses (HPVs) comprise a large family of small, nonenveloped DNA viruses with transforming potential. HPVs selectively infect basal keratinocytes of stratified skin and mucosal epithelia and persist, mostly without clinical symptoms, in virtually every part of the human skin. The biological costs of HPV persistence range from benign papilloma and genital warts over preneoplastic lesions to anogenital or oropharyngeal cancers (1). In fact, infection by the so-called "high-risk" HPV causes about 5% of all human cancers (2). Of these, cervical cancers are the most prevalent. However, HPV-associated oropharyngea...

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Active center differences between cathepsins L and B: The S₁ binding region

Cited by 64 publications

References 25 publications

The Detection, Quantification and Partial Characterisation of Cathepsin B-Like Activity in Human Pathological Synovial Fluids

The Detection, Quantification and Partial Characterisation of Cathepsin B-Like Activity in Human Pathological Synovial Fluids

Characterisation of a cysteine protease from bloodstream forms of Trypanosoma congolense

Kallikrein-8 Proteolytically Processes Human Papillomaviruses in the Extracellular Space To Facilitate Entry into Host Cells

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Active center differences between cathepsins L and B: The S1 binding region

Cited by 64 publications

References 25 publications

The Detection, Quantification and Partial Characterisation of Cathepsin B-Like Activity in Human Pathological Synovial Fluids

The Detection, Quantification and Partial Characterisation of Cathepsin B-Like Activity in Human Pathological Synovial Fluids

Characterisation of a cysteine protease from bloodstream forms of Trypanosoma congolense

Kallikrein-8 Proteolytically Processes Human Papillomaviruses in the Extracellular Space To Facilitate Entry into Host Cells

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Active center differences between cathepsins L and B: The S₁ binding region