Biochemie natürlicher Proteinase‐Inhibitoren

Tschesche, Harald

doi:10.1002/ange.19740860104

Cited by 45 publications

(14 citation statements)

References 172 publications

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“…Modified inhibitors become inactivated upon removal of their reactive4 te residue PI by carboxypeptidases [2,3,5], since the association constant for complex formation is drastically lowered.…”

Section: Lysine-15 Repkdcement In Bovine Inhibitormentioning

confidence: 99%

“…No cleavage of this particular bond has so far been detected under conditions which in other classes of proteinase inhibitors lead to a partial proteolytic cleavage [2,3]. However, the modified trypsin-kallikrein inhibitor (Kunitz) could bc prepared in a series of chemical and enzymic reactions [8,10,11].…”

Section: Lysine-15 Repkdcement In Bovine Inhibitormentioning

confidence: 99%

“…Generally this bond is subject to a limited proteolytic cleavage by catalytic amounts of the inhibited enzymes forming stable stoichiometric complexes with the inhibitor [2,3]. In a true thermodynamic equilibrium modified inhibitors are formed with their -P,-P;-peptide bond hydrolyzed [2-41.…”

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confidence: 99%

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Replacement of Lysine by Arginine, Phenylalanine and Tryptophan in the Reactive Site of the Bovine Trypsin‐Kallikrein Inhibitor (Kunitz) and Change of the Inhibitory Properties

Jering¹,

Tschesche²

1976

European Journal of Biochemistry

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The reactive-site sequence of a proteinase inhibitor can be written as . . . -P,-P2-P,-P;-P;-P;-. . . , where -P,-P;-denotes the reactive site. Three semisynthetic homologues have been synthesized of the bovine trypsin-kallikreinm inhibitor (Kunitz) with either arginine, phenylalanine or tryptophan in place of the reactive-site residue PI, lysine-15. These homologues correspond to gene products after mutation of the lysine 15 DNA codon to an arginine, phenylalanine or tryptophan DNA codon.Starting from native (virgin) inhibitor, reactive-site hydrolyzed, still active (modified) inhibitor was prepared by chemical and enzymic reactions. Modified inhibitor was then converted into inactive des-Lys15-inhibitor by reaction with carboxypeptidase B. Inactive des-Lys"-inhibitor was reactivated by enzymic replacement of the P, residue according to Leary and Laskowski, Jr. The introduction of arginine was catalyzed by an inverse reaction with carboxypeptidase B, while phenylalanine or tryptophan were replaced by carboxypeptidase A. The reactivated semisynthetic inhibitors were trapped by complex formation with either trypsin or chymotrypsin. The enzyme . inhibitor complexes were subjected to kinetic-control dissociation, and the semisynthetic virgin inhibitors were isolated.The inhibitory properties of the semisynthetic inhibitors have been investigated against bovine trypsin and chymotrypsin and against porcine pancreatic kallikrein@ and plasmin. The homologues with either lysine or arginine in the P, position are equally good inhibitors of trypsin, plasmin and kallikrein. The Arg'5-homologue is a slightly more effective kallikrein inhibitor than the Lys"-inhibitor. The semisynthetic phenylalanine and tryptophan homologues, however, are weak inhibitors of trypsin and still weaker inhibitors of kallikrein, but are excellent inhibitors of chymotrypsin. Their association constant with chymotrypsin is at least ten times higher than that of native Lys"-inhibitor. A dramatic specificity change is observed with the phenylalanine and tryptophan homologues, which in contrast to the native inhibitor do not at all inhibit porcine plasmin. Thus, the nature of the P, residue strongly influences the primary inhibitory specificity of the bovine inhibitor (Kunitz). Phe15 and Trp15-inhibitor, virgin trypsin-kallikrein inhibitor homologues with arginine, phenylalanine or tryptophan respectively in place of the Lys-15 reactive-site residue PI ; des-Lys"-inhibitor has the active-site Lys-15 removed by carboxypetidase B. Bz-Arg-Nan, N-bcnzoyl-~~-arginine-4-nitroanilide; Bz-Arg-OEt, N-benzoyl-arginine ethyl ester; Suc-Phe-Nan, N-(3-carboxypropionyl)-phenylalanine-4-nitroanilide; NphGBz, 4-nitrophenyl4'-guanidinobenzoate hydrochloride.

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Section: Lysine-15 Repkdcement In Bovine Inhibitormentioning

confidence: 99%

Section: Lysine-15 Repkdcement In Bovine Inhibitormentioning

confidence: 99%

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Replacement of Lysine by Arginine, Phenylalanine and Tryptophan in the Reactive Site of the Bovine Trypsin‐Kallikrein Inhibitor (Kunitz) and Change of the Inhibitory Properties

Jering¹,

Tschesche²

1976

European Journal of Biochemistry

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“…The conditions for this treatment were adapted from the data for proteinase-inhibitor dissociation outlined by Tschesche [16]. The unmasking was effected by equilibrating the protein solutions (in water or buffers) with about 3 -5 vol.…”

Section: Unmasking Of' Cathepsin R In Rihosonzal Prorein Prepurutionsmentioning

confidence: 99%

The Ribosomal Serine Proteinase, Cathepsin R

Langner¹,

Kirschke²,

Bohley³

et al. 1982

European Journal of Biochemistry

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Ribosomes have been shown to contain a proteolytic activity, characterized as an endopeptidase with serine in the active center. The enzyme has been given the name cathepsin R, following the recommendations of Barrett et al. (in a publication from the Cold Spring Harbor Laboratory, New York) for naming new proteinases. The present paper contains evidence that cathepsin R in rat liver ribosomes is present in a cryptic form. Upon dissociation of ribosomes to subunits (and to minor extent also by 0.5 M KCI washes). the cryptic proteinase is released. Activation of the released cathepsin R is effected by equilibration with 2 M NaCI/O.OS M sodium acetate, pH 4.8. The molecular weight of free cathepsin R is 25000-30000.

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“…The proteinaceous protease inhibitors of animal and plant origins have been extensively studied in the past decades [1][2][3]. Yet, little attention has been paid to the occurence of such inhibitors in microorganisms.…”

Section: Introductionmentioning

confidence: 99%