Thiol-dependent metallo-endopeptidase characteristics of Pz-peptidase in rat and rabbit

Tisljar, Ursula; Barrett, Alan J.

doi:10.1042/bj2670531

Cited by 36 publications

(36 citation statements)

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“…Thimet oligopeptidase (EP 24.15) was purified from rat testis cytosol and rat liver mitochondria by standard methods (Heidrich et al, 1973;Tisljar and Barrett, 1990b). For biochemical comparison of the enzyme with proteinase yscD, the yeast enzyme was purified as described for rat testis EP 24.15 (Tisljar and Barrett, 1990b).…”

Section: Enzyme Purificationmentioning

confidence: 99%

“…For biochemical comparison of the enzyme with proteinase yscD, the yeast enzyme was purified as described for rat testis EP 24.15 (Tisljar and Barrett, 1990b). Otherwise proteinase yscD was purified from strain ABYSl by the procedure described by Achstetter et al (1985).…”

Section: Enzyme Purificationmentioning

confidence: 99%

“…Through sequencing data of the gene and through biochemical data we provide evidence that proteinase yscD is the equivalent of mammalian metalloendopeptidase (EP 24.15; Buchler, 1992) also known as Pz-peptidase (Pz, phenylazobenzoxycarbonyl ; and endo-oligopeptidase A (Harper and Gross, 1970; Coelho et al, 1981 ;. Due to its characteristics, cleaving only oligopeptides and its activity being de- (1987) this study this study this study this study Yanisch-Perron et al (1985) Miller (1987) pendent on metal ions (Znz+) and sensitive to thiol-modifying agents (Orlowski et al, 1983;Orlowski et al, 1988;Orlowski et al, 1989), the enzyme had been named thimet oligopeptidase recently (Tisljar and Barrett, 1990b). The mammalian enzyme is present in the cytoplasm and mitochondria of cells (Heidrich et al, 1973;Tisljar and Barrett, 1990a).…”

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Proteinase yscD (oligopeptidase yscD)

Büchler

Tisljar

Wolf

1994

European Journal of Biochemistry

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The yeast PRDl gene, encoding proteinase yscD, was cloned by complementation of the prdl-6 point mutation. Sequencing of the gene revealed an open reading frame of 2.136 kb, encoding a protein of 712 amino acids with a calculated molecular mass of 81.8 kDa. The sequence HEGLG beginning at residue 501 represents the HEXXH motif, unique for the zinc metallo-peptidases. (RS)-carboxy-3-phenylpropyl]-Ala-Ala-Phe-p-aminobenzoic acid, a compound designed as specific inhibitor of EP 24.15, is also a strong inihibitor of the yeast enzyme. Proteinase yscD is a nonvacuolar enzyme. 3-5% of the total enzyme activity can be detected in the intermembrane space of mitochondria. In a mutant carrying a deletion of the PRDl gene no proteinase yscD activity is detectable in the cytoplasm and in mitochondria of these cells. They do not show any grossly altered phenotype but exhibit a decrease in the intracellular degradation of peptides. This suggests a function of proteinase yscD in the late stages of protein degradation.Proteolysis is a multifunctional, essential principle in in-

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Section: Enzyme Purificationmentioning

confidence: 99%

Section: Enzyme Purificationmentioning

confidence: 99%

mentioning

confidence: 99%

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Proteinase yscD (oligopeptidase yscD)

Büchler

Tisljar

Wolf

1994

European Journal of Biochemistry

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“…After a debris spin of 5 min at 1000 x g, the supernatant was centrifuged for 20 min at 10 000 x g. The resulting mitochondrial pellet was washed in the sucrose/Tris buffer and resuspended in 50 mM Tris/HCl, pH 8.0, 2% (w/v) Triton X-100. After 30 min at 4°C the preparation was centrifuged for 20 min at 10 000 x g and the supernatant was run on columns of DEAEcellulose, Sephacryl S-200 HR, Mono Q and Mono P as previously described for the rat testis enzyme [3], except that 0.2% Brij 35 was included in all buffers after the DEAE-cellulose step.…”

Section: Enzyme Purificationmentioning

confidence: 99%

“…Thimet peptidase (EC 3.4.24.15), previously known as Pz-peptidase, endo-ohgopeptidase A [l] and soluble metallo-endopeptidase [2] is a thiol-dependent metalloendopeptidase [3] that acts on oligopeptides [4,5]. Most published reports have concerned the cytosolic form of the enzyme, but Heidrich et al [6] discovered a 'Pzpeptidase' in highly purified mitochondria from rat liver.…”

Section: Introductionmentioning

confidence: 99%

A distinct thimet peptidase from rat liver mitochondria

Tisljar

Barrett

1990

FEBS Letters

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Thimet peptidase has been purified from rat liver mitochondria and found to share the characteristics of a thiol-dependent metallo-endopeptidase previously described for an enzyme in the cytosolic fraction from rat testis: inhibition by EDTA, reactivation by Zn2+, requirement of dithiothreitol for maximal and stable activity, and inhibition by N-ethylmaleimide. The K, for inhibition by N-[1-(RS)-carboxy-3-phenylpropyl]-Ala-Ala-Phe-paminobenzoic acid is 2.6 PM, 100-fold higher than the value for the cytosolic form. The mitochondrial form is not inhibited by antisera against the cytosolic form, and the two forms of the enzyme show quantitative differences in substrate specificity. The name thimet peptidase II is suggested for the enzyme from rat mitochondria.

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Regulators of the neuropeptide‐degrading enzyme, EC 3.4.24.15 (thimet oligopeptidase), in cerebrospinal fluid

Shrimpton

Wolfson

Smith

et al. 2003

J of Neuroscience Research

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Endopeptidase EC 3.4.24.15 (EP 24.15; thimet oligopeptidase) is a soluble metalloendopeptidase implicated in the metabolism of a number of neuropeptides, including neurotensin, gonadotropin-releasing hormone, and opioid peptides. We have shown previously that thiol reducing agents, such as dithiothreitol, activate EP 24.15 by mediating the conversion of inactive multimeric forms to active monomers and that this conversion involves the disruption of intermolecular disulfide bonds involving cysteine residues 246, 248, and 253. We have identified two components of cerebrospinal fluid that activate recombinant EP 24.15, but have no effect on a thiol-independent cysteine mutant form of the enzyme. The low molecular weight (<10 kDa) component co-elutes with glutathione by reversed-phase HPLC, whereas the high molecular weight component (>50 kDa) is sensitive to digestion with trypsin, suggesting it is proteinaceous in nature. These results suggest that EP 24.15 activity in the brain may be modulated by factors released into cerebrospinal fluid.

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Thiol-dependent metallo-endopeptidase characteristics of Pz-peptidase in rat and rabbit

Cited by 36 publications

References 12 publications

Proteinase yscD (oligopeptidase yscD)

Proteinase yscD (oligopeptidase yscD)

A distinct thimet peptidase from rat liver mitochondria

Regulators of the neuropeptide‐degrading enzyme, EC 3.4.24.15 (thimet oligopeptidase), in cerebrospinal fluid

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