HOME HELP FEEDBACK SUBSCRIPTIONS ARCHIVE SEARCH TABLE OF CONTENTS		QUICK SEARCH:   [advanced] Author: Keyword(s): Year:  Vol:  Page:

This Article Full Text Full Text (PDF) All Versions of this Article: ps.04759104v1 13/9/2388    most recent Alert me when this article is cited Alert me if a correction is posted Citation Map Services Email this article to a friend Similar articles in this journal Similar articles in PubMed Alert me to new issues of the journal Download to citation manager Citing Articles Citing Articles via HighWire Citing Articles via Google Scholar Google Scholar Articles by Machczynski, M. C. Articles by Canters, G. W. Search for Related Content PubMed PubMed Citation Articles by Machczynski, M. C. Articles by Canters, G. W. Social Bookmarking                   What's this?

Characterization of SLAC: A small laccase from Streptomyces coelicolor with unprecedented activity

¹ Leiden Institute of Chemistry, Gorlaeus Laboratories, Leiden University, 2333 CC, Leiden, The Netherlands
² Department of Biochemistry, Physiology, and Microbiology, University of Gent, B-9000 Gent, Belgium

(RECEIVED March 23, 2004; FINAL REVISION May 18, 2004; ACCEPTED May 18, 2004)

Laccases and other four-copper oxidases are usually constructed of three domains: Domains one and three house the copper sites, and the second domain often helps form a substrate-binding cleft. In contrast to this arrangement, the genome of Streptomyces coelicolor was found to encode a small, four-copper oxidase that lacks the second domain. This protein is representative of a new family of enzymes—the two-domain laccases. Disruption of the corresponding gene abrogates laccase activity in the growth media. We have recombinantly expressed this enzyme, called SLAC, in Escherichia coli and characterized it. The enzyme binds four copper ions/monomer, and UV-visible absorption and EPR measurements confirm that the conserved type 1 copper site and trinuclear cluster are intact. We also report the first known paramagnetic NMR spectrum for the trinuclear copper cluster of a protein from the laccase family. The enzyme is highly stable, retaining activity as a dimer in denaturing gels after boiling and SDS treatment. The activity of the enzyme against 2,6-dimethoxyphenol (DMP) peaks at an unprecedentedly high pH (9.4), whereas the activity against ferrocyanide decreases with pH. SLAC binds negatively charged substrates more tightly than positively charged or uncharged molecules.

Keywords: laccase; NMR; activity; multicopper; oxidase; copper; PPO

Article published online ahead of print. Article and publication date are at http://www.proteinscience.org/cgi/doi/10.1110/ps.04759104.

Reprint requests to: Gerard W. Canters, Leiden University, Einsteinweg 55, P.O. Box 9502, 2300 RA Leiden, The Netherlands; e-mail: canters{at}chem.leidenuniv.nl; fax: +31-71-527-4349.

CiteULike    Connotea    Del.icio.us    Digg    Reddit    Technorati    What's this?

This article has been cited by other articles:

				I. R. Wheeldon, J. W. Gallaway, S. C. Barton, and S. Banta Bioelectrocatalytic hydrogels from electron-conducting metallopolypeptides coassembled with bifunctional enzymatic building blocks PNAS, October 7, 2008; 105(40): 15275 - 15280. [Abstract] [Full Text] [PDF]

				A. Beloqui, M. Pita, J. Polaina, A. Martinez-Arias, O. V. Golyshina, M. Zumarraga, M. M. Yakimov, H. Garcia-Arellano, M. Alcalde, V. M. Fernandez, et al. Novel Polyphenol Oxidase Mined from a Metagenome Expression Library of Bovine Rumen: BIOCHEMICAL PROPERTIES, STRUCTURAL ANALYSIS, AND PHYLOGENETIC RELATIONSHIPS J. Biol. Chem., August 11, 2006; 281(32): 22933 - 22942. [Abstract] [Full Text] [PDF]