期刊
ACTA CRYSTALLOGRAPHICA SECTION F-STRUCTURAL BIOLOGY COMMUNICATIONS
卷 74, 期 -, 页码 496-505出版社
INT UNION CRYSTALLOGRAPHY
DOI: 10.1107/S2053230X18006842
关键词
endoglucanase; Ruminiclostridium thermocellum; Clostridium; metal ion; oligosaccharides; enzymes; carbohydrates; bio-inorganic; 2-oxohistidine
资金
- Biotechnology and Biological Sciences Research Council [BB/L021633/1]
- BBSRC [BB/L021633/1, BB/R007705/1] Funding Source: UKRI
The recent discovery of 'lytic' polysaccharide monooxygenases, copper-dependent enzymes for biomass degradation, has provided new impetus for the analysis of unusual metal-ion sites in carbohydrate-active enzymes. In this context, the CAZY family GH124 endoglucanase from Ruminiclostridium thermocellum contains an unusual metal-ion site, which was originally modelled as a Ca-2(+) site but features aspartic acid, asparagine and two histidine imidazoles as coordinating residues, which are more consistent with a transition-metal binding environment. It was sought to analyse whether the GH124 metal-ion site might accommodate other metals. It is demonstrated through thermal unfolding experiments that this metal-ion site can accommodate a range of transition metals (Fe-2(+), Cu-2(+), Mn-2(+) and Ni-2(+)), whilst the three-dimensional structure and mass spectrometry show that one of the histidines is partially covalently modified and is present as a 2-oxohistidine residue; a feature that is rarely observed but that is believed to be involved in an 'off-switch' to transition-metal binding. Atomic resolution (<1.1 angstrom) complexes define the metal-ion site and also reveal the binding of an unusual fructosylated oligosaccharide, which was presumably present as a contaminant in the cellohexaose used for crystallization. Although it has not been possible to detect a biological role for the unusual metal-ion site, this work highlights the need to study some of the many metal-ion sites in carbohydrate-active enzymes that have long been overlooked or previously mis-assigned.
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