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Illuminating structure and acyl donor sites of a physiological transglutaminase substrate from Streptomyces mobaraensis.

Juettner, Norbert E. and Schmelz, Stefan and Bogen, Jan P. and Happel, Dominic and Fessner, Wolf-Dieter and Pfeifer, Felicitas and Fuchsbauer, Hans-Lothar and Scrima, Andrea (2018):
Illuminating structure and acyl donor sites of a physiological transglutaminase substrate from Streptomyces mobaraensis.
In: Protein science : a publication of the Protein Society, pp. 910-922, 27, (5), ISSN 1469-896X, [Article]

Abstract

Transglutaminase from Streptomyces mobaraensis (MTG) has become a powerful tool to covalently and highly specifically link functional amines to glutamine donor sites of therapeutic proteins. However, details regarding the mechanism of substrate recognition and interaction of the enzyme with proteinaceous substrates still remain mostly elusive. We have determined the crystal structure of the Streptomyces papain inhibitory protein (SPI), a substrate of MTG, to study the influence of various substrate amino acids on positioning glutamine to the active site of MTG. SPIexhibits a rigid, thermo-resistant double-psi-beta-barrel fold that is stabilized by two cysteine bridges. Incorporation of biotin cadaverine identified Gln-6 as the only amine acceptor site on SPIaccessible for MTG. Substitution of Lys-7 demonstrated that small and hydrophobic residues in close proximity to Gln-6 favour MTG-mediated modification and are likely to facilitate introduction of the substrate into the front vestibule of MTG. Moreover, exchange of various surface residues of SPIfor arginine and glutamate/aspartate outside the glutamine donor region influences the efficiency of modification by MTG. These results suggest the occurrence of charged contact areas between MTG and the acyl donor substrates beyond the front vestibule, and pave the way for protein engineering approaches to improve the properties of artificial MTG-substrates used in biomedical applications. This article is protected by copyright. All rights reserved.

Item Type: Article
Erschienen: 2018
Creators: Juettner, Norbert E. and Schmelz, Stefan and Bogen, Jan P. and Happel, Dominic and Fessner, Wolf-Dieter and Pfeifer, Felicitas and Fuchsbauer, Hans-Lothar and Scrima, Andrea
Title: Illuminating structure and acyl donor sites of a physiological transglutaminase substrate from Streptomyces mobaraensis.
Language: English
Abstract:

Transglutaminase from Streptomyces mobaraensis (MTG) has become a powerful tool to covalently and highly specifically link functional amines to glutamine donor sites of therapeutic proteins. However, details regarding the mechanism of substrate recognition and interaction of the enzyme with proteinaceous substrates still remain mostly elusive. We have determined the crystal structure of the Streptomyces papain inhibitory protein (SPI), a substrate of MTG, to study the influence of various substrate amino acids on positioning glutamine to the active site of MTG. SPIexhibits a rigid, thermo-resistant double-psi-beta-barrel fold that is stabilized by two cysteine bridges. Incorporation of biotin cadaverine identified Gln-6 as the only amine acceptor site on SPIaccessible for MTG. Substitution of Lys-7 demonstrated that small and hydrophobic residues in close proximity to Gln-6 favour MTG-mediated modification and are likely to facilitate introduction of the substrate into the front vestibule of MTG. Moreover, exchange of various surface residues of SPIfor arginine and glutamate/aspartate outside the glutamine donor region influences the efficiency of modification by MTG. These results suggest the occurrence of charged contact areas between MTG and the acyl donor substrates beyond the front vestibule, and pave the way for protein engineering approaches to improve the properties of artificial MTG-substrates used in biomedical applications. This article is protected by copyright. All rights reserved.

Journal or Publication Title: Protein science : a publication of the Protein Society
Volume: 27
Number: 5
Divisions: 10 Department of Biology
10 Department of Biology > Microbiology and Archaea
Date Deposited: 20 Feb 2018 08:56
Identification Number: pmid:29430769
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