Expression and partial characterization of a cathepsin B-like enzyme (Sm31) and a proposed ‘haemoglobinase’ (Sm32) from <i>Schistosoma mansoni</i>

Götz, Bernhard; Klinkert, Mo‐Quen

doi:10.1042/bj2900801

Cited by 49 publications

(21 citation statements)

References 29 publications

(24 reference statements)

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“…VPEs are classified as a novel family of cysteine proteinase. The plant VPEs exhibit homology to a putative cysteine proteinase, Sm32, of the human parasite Schistosoma mansoni [7,21]. However, the physiological function of the Schistosoma enzyme is still unknown.…”

Section: Discussionmentioning

confidence: 99%

Homologues of a vacuolar processing enzyme that are expressed in different organs in Arabidopsis thaliana

1995

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Vacuolar processing enzymes (VPEs) are responsible for the maturation of seed proteins. These processing enzymes belong to a novel group of cysteine proteinases with molecular masses of 37 to 39 kDa. We isolated two genes of VPEs from a genomic library of Arabidopsis. The gene products were designated alpha-VPE and beta-VPE, and they were 56% identical in terms of amino acid sequence. The amino acid sequences of alpha-VPE and beta-VPE were also 55% and 67% identical to that of castor bean VPE, respectively. The gene for alpha-VPE had 7 introns, while that of beta-VPE had 8 introns. Northern blot analysis revealed that alpha-VPE is expressed in rosette leaves, cauline leaves and stems of Arabidopsis, while beta-VPE is predominantly expressed in the flowers and buds. Neither alpha-VPE nor beta-VPE is expressed in the siliques. This result strongly suggests that the isolated genes encode isozymes of VPE that are specific to vegetative organs.

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Section: Discussionmentioning

confidence: 99%

Homologues of a vacuolar processing enzyme that are expressed in different organs in Arabidopsis thaliana

1995

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“…Clostripain probably does not form specific S' subsite loops as found in common serine proteases (Schellenberger et al, 1993a). Gilles et al (1984), and Gotz and Klinkert (1993) stated that the primary structure of clostripain is not homologous with either other cysteine proteases or with any other known protein structure. The outstanding behaviour in forming peptide bonds with proline in the P: position underlines how extraordinary this enzyme is and indicates that hydrogen bonds are not significantly involved in the binding of the nucleophile and the enzyme subsite.…”

Section: Discussionmentioning

confidence: 99%

The specificity of clostripain from Clostridium histolyticum

Ullmann¹,

Jakubke²

1994

European Journal of Biochemistry

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The S' subsite specificity of clostripain from Clostridium histolyticum was investigated by acyl transfer to libraries of amino acid amides, Ala-Xaa dipeptides, proline derivatives and pentapeptides using Nu-benzoyl-L-arginine ethyl ester as acyl donor. A pentapeptide library consisting of 29 pentapeptides with general structure Xaa-Ala-Ala-Ala-Gly, Ala-Xaa-Ala-Ala-Gly and Ala-Ala-XaaAla-Gly, where Xaa represents Gly, Ala, Pro, Leu, Phe, Asp, Glu, Arg and Lys, was prepared by solid-phase synthesis. The data analysis was performed by HPLC and evaluated by statistical algorithms. The nucleophile efficiency covers a range of more than three orders of magnitude. In the P; position, low specificity for amino acid amides and Xaa-(Ala),-Gly peptides was found, however, in the Pi position, positively charged amino acid residues are strongly preferred. The negatively charged side chains of aspartic acid and glutamic acid in the P: and Pi positions, respectively, show only poor nucleophilic behaviour. In the case of these amino acids, the S'-P' interactions depend significantly on their position of these residues in the peptide chain of the nucleophile. The transfer of aspartic acid and glutamic acid from P; -Pi increases the nucleophile efficiency by approximately two orders of magnitude. The aromatic side chains are not well accepted, especially in the case of PiPhe. Surprisingly, P:Gly leads to effective P'-S' interactions. However, the opposite result was obtained for PiGly. The high efficiency for Gly-NH, does not fit with the hydrophobicity structure/ activity relationships. In most cases, peptide chain elongation does not improve the nucleophile efficiency. The effective interaction of D-Leu-NH, with the S' subsite of clostripain emphasizes the fact that the nucleophile stereospecificity is not restricted to L-amino acids. The results with proline derivatives indicate remarkably different specificities of the S' binding site which can only be explained by conformational restraints. A positive cooperativity between P:Pro and PiGly and a negative cooperativity between Pi Pro and PiPhe was observed. The arrangement of three proline residues next to each other represents a favourable conformation for effective enzyme-nucleophile interactions.The cysteine protease clostripain (Clostridium histolyticum proteinase B) from the culture filtrate of C. histolyticum is well known for its restricted substrate specificity, the cleavage of Arg-Xaa bonds, including proline in the P; position (subsite nomenclature of Schechter and Berger, 1967) (Ogle andTytell, 1953 ;Labouesse and Gros, 1960; Mitchell, 1964). For this reason this enzyme is important both in sequence analysis and in enzymic peptide synthesis, because proline is normally not well accepted in the P: position by the majority of proteases. Furthermore, a precise characterization of clostripain specificity is important if selective inhibitors are to be designed.Although S subsites have been studied in detail (Cole et al., 1971;Porter et al., 1971;Gilles et al., 1979;

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“…It is likely that proteolysis and/or autolysis around the asparagine residue induces a conformational change in the inactive precursor and then generates an active vacuolar processing enzyme in vivo and in E. coli transformed with pVPEdel. Gotz and Klinkert (1993) reported that insect cells expressing the complete cDNA sequence of the putative cysteine proteinase of the human parasite S. mansoni has no proteolytib activity on hemoglobin. They suggested that the expressed protein is not further cleaved to produce an active form.…”

Section: Discussionmentioning

confidence: 99%

Molecular characterization of a vacuolar processing enzyme related to a putative cysteine proteinase of Schistosoma mansoni.

1993

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Proproteins of various vacuolar proteins are post-translationally processed into mature forms by the action of a unique vacuolar processing enzyme. If such a processing enzyme is transported to vacuoles together with proprotein substrates, the enzyme must be a latent form. lmmunocytochemical localization of a vacuolar processing enzyme, a 37-kD cysteine proteinase, in the endosperm of maturing castor bean seeds places the enzyme in the vacuolar matrix, where a variety of proproteins is also present. To characterize a molecular structure of vacuolar processing enzyme, we isolated a cDNA for the enzyme. Deduced primary structure of a 55-kD precursor is 33% identical to a putative cysteine proteinase of the human parasite Schistosoma mansoni. The precursor is composed of a signal peptide, a 37-kD active processing enzyme domain, and a propeptide fragment. Although the precursor expressed in Escherichia coli has no vacuolar processing activity, a 36-kD immunopositive protein expressed in E. coli is active. These results suggest that the activation of the vacuolar processing enzyme requires proteolytic cleavage of a 1CkD C-terminal propeptide fragment of the precursor.

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Expression and partial characterization of a cathepsin B-like enzyme (Sm31) and a proposed ‘haemoglobinase’ (Sm32) from Schistosoma mansoni

Cited by 49 publications

References 29 publications

Homologues of a vacuolar processing enzyme that are expressed in different organs in Arabidopsis thaliana

Homologues of a vacuolar processing enzyme that are expressed in different organs in Arabidopsis thaliana

The specificity of clostripain from Clostridium histolyticum

Molecular characterization of a vacuolar processing enzyme related to a putative cysteine proteinase of Schistosoma mansoni.

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