Specificity of prohormone convertase 2 on proenkephalin and proenkephalin-related substrates

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dc.contributor.author Johanning, K.
dc.contributor.author Juliano, Maria Aparecida [UNIFESP]
dc.contributor.author Juliano, Luiz [UNIFESP]
dc.contributor.author Lazure, C.
dc.contributor.author Lamango, N. S.
dc.contributor.author Steiner, D. F.
dc.contributor.author Lindberg, I
dc.date.accessioned 2016-01-24T12:30:38Z
dc.date.available 2016-01-24T12:30:38Z
dc.date.issued 1998-08-28
dc.identifier http://dx.doi.org/10.1074/jbc.273.35.22672
dc.identifier.citation Journal of Biological Chemistry. Bethesda: Amer Soc Biochemistry Molecular Biology Inc, v. 273, n. 35, p. 22672-22680, 1998.
dc.identifier.issn 0021-9258
dc.identifier.uri http://repositorio.unifesp.br/handle/11600/25941
dc.description.abstract In the central and peripheral nervous systems, the neuropeptide precursor proenkephalin must be endoproteolytically cleaved by enzymes known as prohormone convertases 1 and 2 (PC1 and PC2) to generate opioid-active enkephalins. in this study, we have investigated the specificity of recombinant mouse PC2 for proenkephalin-related internally quenched (IQ) peptides, for methylcoumarin amide-based fluorogenic peptides, and for recombinant rat proenkephalin. IQ peptides exhibited specificity constants (k(cat)/K-m) between 9.4 x 10(4) M-1 s(-1)(Abz-Val-Pro-Arg-Met-Glu-Lys-Arg-Tyr-Gly-Gly-Phe-Met-Gln-EDDnp; where Abz is ortho-aminobenzoic acid and EDDnp is N-(2,4-dinitrophenyl)ethylenediamine)) and 0.24 x 10(4) M-1 s(-1) (Abz-Tyr-Gly-Gly-Phe-Met-Arg-Arg-Val-Gly-Arg-Pro-Glu-EDDnp), with the peptide B to Met-enk-Arg-Phe cleavage preferred (Met-enk is met-enkephalin). Fluorogenic substrates with P1, P2, and P4 basic amino acids were hydrolyzed with specificity constants ranging between 2.0 x 10(3) M-1 s(-1) (Ac-Orn-Ser-Lys-Arg-MCA; where MCA is methylcoumarin amide) and 1.8 x 10(4) M-1 s(-1) (<Glu-Arg-Thr-Lys-Arg-MCA; where <Glu is pyroglutamic acid). Substrates containing only a single basic residue were not appreciably hydrolyzed, and substrates lacking a P4 Arg exhibited k(cat) of less than 0.05 s(-1). Substitution of ornithine for Lys at the P4 position did not significantly affect the k(cat) but increased the K-m 2-fold. Data from both sets of fluorogenic substrates supported the contribution of a P4 Arg to PC2 preference. Analysis of proenkephalin reaction products using immunoblotting and gel permeation chromatography demonstrated that PC2 can directly cleave proenkephalin and that the generation of small opioid peptides from intermediates is mediated almost entirely by PC2 rather than by PC1, These results are in accord with the analysis of PC2 knock-out brains, in which the amounts of three mature enkephalins were depleted by more than three-quarters. en
dc.format.extent 22672-22680
dc.language.iso eng
dc.publisher Amer Soc Biochemistry Molecular Biology Inc
dc.relation.ispartof Journal of Biological Chemistry
dc.rights Acesso aberto
dc.title Specificity of prohormone convertase 2 on proenkephalin and proenkephalin-related substrates en
dc.type Artigo
dc.contributor.institution Louisiana State Univ
dc.contributor.institution Universidade Federal de São Paulo (UNIFESP)
dc.contributor.institution Inst Rech Clin Montreal
dc.contributor.institution Univ Chicago
dc.description.affiliation Louisiana State Univ, Med Ctr, Sch Med, Dept Biochem & Mol Biol, New Orleans, LA 70112 USA
dc.description.affiliation Escola Paulista Med, Dept Biophys, BR-04044020 São Paulo, Brazil
dc.description.affiliation Inst Rech Clin Montreal, Neuropeptide Struct & Metab Lab, Quebec City, PQ 2W 1R7, Canada
dc.description.affiliation Univ Chicago, Sch Med, Howard Hughes Med Inst, Chicago, IL 60637 USA
dc.description.affiliation Univ Chicago, Sch Med, Dept Biochem, Chicago, IL 60637 USA
dc.description.affiliationUnifesp Escola Paulista Med, Dept Biophys, BR-04044020 São Paulo, Brazil
dc.identifier.doi 10.1074/jbc.273.35.22672
dc.description.source Web of Science
dc.identifier.wos WOS:000075616600073

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