pH stability of the stromelysin-1 catalytic domain and its mechanism of interaction with a glyoxal inhibitor

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Show simple item record Howe, Nicole Ceruso, Mariangela Spink, Edward Malthouse, J.Paul G. 2011-07-19T14:12:14Z 2011-07-19T14:12:14Z 2011 Elsevier B.V. en 2011-10
dc.identifier.citation Biochimica et Biophysica Acta (BBA) - Proteins & Proteomics en
dc.identifier.issn 1570-9639
dc.description.abstract The Stromelysin-1 catalytic domain83-247 (SCD) is stable for at least 16 hours at pHs 6.0-8.4. At pHs 5.0 and 9.0 there is exponential irreversible denaturation with half lives of 38 and 68 min respectively. At pHs 4.5 and 10.0 irreversible denaturation is biphasic. At 25°C, C-terminal truncation of stromelysin-1 decreases the stability of the stromelysin-1 catalytic domain at pH values > 8.4 and < 6.0. We describe the conversion of the carboxylate group of (βR)-β-[[[(1S)-1-[[[(1S)-2-Methoxy-1-phenylethyl]amino]carbonyl]-2,2-dimethylpropyl]amino]carbonyl]-2-methyl-[1,1'-biphenyl]-4-hexanoic acid (UK-370106-COOH) a potent inhibitor of the metalloprotease stromelysin-1 to a glyoxal group (UK-370106-CO13CHO). At pH 5.5 - 6.5 the glyoxal inhibitor is a potent inhibitor of stromelysin-1 (Ki = ~1 μM). The aldehyde carbon of the glyoxal inhibitor was enriched with carbon-13 and using Carbon-13 NMR we show that the glyoxal aldehyde carbon is fully hydrated when it is in aqueous solutions (90.4 ppm) and also when it is bound to SCD (~92.0 ppm). We conclude that the hemiacetal hydroxyl groups of the glyoxal inhibitor are not ionised when the glyoxal inhibitor is bound to SCD. The free enzyme pKa values associated with inhibitor binding were 5.9 and 6.2. The formation and breakdown of the signal at ~92 ppm due to the bound UK-370106-CO13CHO inhibitor depends on pKa values of 5.8 and 7.8 respectively. No strong hydrogen bonds are present in free SCD or in SCD-inhibitor complexes. We conclude that the inhibitor glyoxal group is not directly coordinated to the catalytic zinc atom of SCD. en
dc.description.sponsorship Science Foundation Ireland en
dc.description.sponsorship Higher Education Authority en
dc.format.extent 1011476 bytes
dc.format.mimetype application/pdf
dc.language.iso en en
dc.publisher Elsevier en
dc.rights This is the author’s version of a work that was accepted for publication in Biochimica et Biophysica Acta (BBA) - Proteins and Proteomics). Changes resulting from the publishing process, such as peer review, editing, corrections, structural formatting, and other quality control mechanisms may not be reflected in this document. Changes may have been made to this work since it was submitted for publication. A definitive version was subsequently published in Biochimica et Biophysica Acta (BBA) - Proteins and Proteomics), DOI: 10.1016/j.bbapap.2011.07.004. en
dc.subject Metalloprotease en
dc.subject Glyoxal inhibitor en
dc.subject pH stability en
dc.subject pKa en
dc.subject Tetrahedral intermediate en
dc.subject.lcsh Metalloproteinases--Inhibitors en
dc.subject.lcsh Metalloproteinases--Stability en
dc.subject.mesh Matrix Metalloproteinase 3 en
dc.subject.mesh Hydrogen-Ion Concentration en
dc.title pH stability of the stromelysin-1 catalytic domain and its mechanism of interaction with a glyoxal inhibitor en
dc.type Journal Article en
dc.internal.availability Full text available en
dc.internal.webversions en
dc.status Peer reviewed en
dc.identifier.volume 1814 en
dc.identifier.issue 10 en
dc.identifier.startpage 1394 en
dc.identifier.endpage 1403 en
dc.identifier.doi 10.1016/j.bbapap.2011.07.004
dc.neeo.contributor Howe|Nicole|aut| en
dc.neeo.contributor Ceruso|Mariangela|aut| en
dc.neeo.contributor Spink|Edward|aut| en
dc.neeo.contributor Malthouse|J.Paul G.|aut| en
dc.description.admin NYP? can't find in journal - AV 13/7/2011 en

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