Bovine Cathepsins S and L: Isolation and Amino Acid Sequences

Dolenc, Iztok; Ritonja, Anka; Čolič, Adrijana; Podobnik, M.; Ogrinc, Tadeja; Türk, Vito

doi:10.1515/bchm3.1992.373.2.407

Cited by 19 publications

(15 citation statements)

References 2 publications

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“…The molecular mass of the enzyme was estimated to be 37 kDa by SDS‐PAGE showing it is slightly larger as compared with that of cathepsin S from bovine 2 (20 kDa), rabbit 21 (30 kDa) and human spleen 3 (24 kDa). In the studies on other carp cathepsins, molecular mass of the single chain of cathepsins B, L and H were 29 kDa, 9 30 kDa 12 and 30 kDa, 25 respectively.…”

Section: Discussionmentioning

confidence: 98%

“…Cathepsin S (EC 3.4.22.27), a lysosomal cysteine protease of endo‐type, was purified from bovine lymph nodes originally 1 and has been purified from bovine, rabbit, rat and human tissues 2–4 . It was found that cathepsin S shares a similar enzymatic property with cathepsin L (EC 3.4.22.15).…”

Section: Introductionmentioning

confidence: 99%

“…They have a high specific activity towards protein substrates, and phylogenetically as well 5–7 . However, cathepsin S differs from cathepsin L because it is a single chain enzyme like papain 2–4 . Furthermore, it differs from other endo‐type lysosomal cysteine proteinases since it is stable and active at neutral pH.…”

Section: Introductionmentioning

confidence: 99%

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Purification and characterization of cathepsin S from hepatopancreas of carp Cyprinus carpio

et al. 2000

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SUMMARY: Cathepsin S was purified from carp hepatopancreas to homogeneity up to 300‐fold. The amino acid sequence of its NH2‐terminus was determined to be V‐P‐D‐A‐M‐D‐W‐Y‐N‐K‐G‐Y‐V‐T‐D‐V‐K‐N‐Q. On the contrary, that of purified cathepsin L from carp hepatopancreas was to be V‐P‐N‐S‐L‐D‐W‐R‐E‐K‐G. Purified cathepsin S consisted of a single chain with 37 kDa estimated by sodium dodecylsulfate–polyacrylamide gel electrophoresis. The enzyme had strong hydrolytic activity toward Z‐Phe‐Arg‐MCA with the pH optimum of 7.0, but this lacked the ability to hydrolyze most of the other MCA substrates. The optimum pH of cathepsin S for protein substrate (carp myosin heavy chain) was also to be pH 7.0. These properties of purified cathepsin S obviously differ from cathepsins B and L. The enzyme activity was totally inhibited by E‐64, leupeptin, 5–5′‐dithiobis (2‐nitro‐benzoic acid) and p‐tosyl‐lys chloromethylketone as well.

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Section: Discussionmentioning

confidence: 98%

Section: Introductionmentioning

confidence: 99%

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Purification and characterization of cathepsin S from hepatopancreas of carp Cyprinus carpio

et al. 2000

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“…Stock solutions of the substrates were prepared in dimethylsulfoxide (Merck, Germany). Human LK, chicken cystatin, and bovine cathepsins B, H , S, and L were purified as described previously (Zvonar et al, 1979;Turk et al, 1983;Muller-Ester1 et al, 1988;Dolenc et al, 1992;PopoviE et al, 1993). Papain (2x crystallized; Sigma, St. Louis, Missouri, USA) was further purified by affinity chromatography (Blumberg et al, 1970); the purified enzyme had a thiol content of 0.96 +.…”

Section: Methodsmentioning

confidence: 99%

High-affinity binding of two molecules of cysteine proteinases to low-molecular-weight kininogen

et al. 1995

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Human low-molecular-weight kininogen (LK) was shown by fluorescence titration to bind two molecules of cathepsins L and S and papain with high affinity. By contrast, binding of a second molecule of cathepsin H was much weaker. The 2: 1 binding stoichiometry was confirmed by titration monitored by loss of enzyme activity and by sedimentation velocity experiments. The kinetics of binding of cathepsins L and S and papain showed the two proteinase binding sites to have association rate constants k,,,,, = 10.7-24.5 x IO6 M" s-I a nd k,,,,, = 0.83-1.4 x lo6 M" s-' . Comparison of these kinetic constants with previous data for intact LK and its separated domains indicate that the faster-binding site is also the tighter-binding site and is present on domain 3, whereas the slowerbinding, lower-affinity site is on domain 2. These results also indicate that there is no appreciable steric hindrance for the binding of proteinases between the two binding sites or from the kininogen light chain.

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“…Ep-475 was from Peptide Research Institute (Osaka, Japan); EDTA and papain (EC 3.4.22.2; 2x crystallized) were from Sigma (St. Louis, USA). Bovine cathepsin S [19] and bovine stefin B [7] were purified using published procedures. Cathepsin S (EC 3.4.22.27) and additionally purified papain [26] were active-site titrated with Ep-475 as described previously [9], whereas stefin B was titrated with active-site titrated papain.…”

Section: Methodsmentioning

confidence: 99%

Kinetics of inhibition of bovine cathepsin S by bovine stefin B

et al. 1994

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The kinetics of the complex formation between bovine cathepsin S and bovine stefin B was studied by conventional and stopped-flow techniques.The inhibition at low inhibitor concentrations was tight and reversible (k,,, = 5.8 x 10' M-' s-l, kdlrr = 4.9 x 10m4 s-' at pH 6.0 and 25"C), whereas at higher inhibitor concentrations it was pseudo-irreversible (k,,, = 6.14 x 10' M-' SC'). The complex was formed directly lacking the fast preequilibrium step with the dissociation equilibrium constant of -8 pM. The competitive nature of inhibition was confirmed. The k,, was found to be pH-Independent between pH 6.0 and 7.5 and decreased at lower or higher pH values in a way that strongly suggests involvement of two ionizable groups in the interaction (pK, = 5.2, pKz = 8.3). The enzyme-substrate interaction seems to be influenced by different ionizable groups (pK, = 4.4, pK, = 7.8).

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Bovine Cathepsins S and L: Isolation and Amino Acid Sequences

Cited by 19 publications

References 2 publications

Purification and characterization of cathepsin S from hepatopancreas of carp Cyprinus carpio

Purification and characterization of cathepsin S from hepatopancreas of carp Cyprinus carpio

High-affinity binding of two molecules of cysteine proteinases to low-molecular-weight kininogen

Kinetics of inhibition of bovine cathepsin S by bovine stefin B

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