title: The binding of zinc ions to Emericella nidulans endo-β-1,4-galactanase is essential for crystal formation.
abstract: A novel Emericella nidulans endo-β-1,4-galactanase (EnGAL) demonstrates a strong capacity to generate high levels of very potent prebiotic oligosaccharides from potato pulp, a by-product of the agricultural potato-starch industry. EnGAL belongs to glycoside hydrolase family 53 and shows high (72.5%) sequence identity to an endo-β-1,4-galactanase from Aspergillus aculeatus. Diffraction data extending to 2.0 Å resolution were collected from a crystal of EnGAL grown from conditions containing 0.2 M zinc acetate. The crystal structure showed a high similarity between EnGAL and other endo-β-1,4-galactanases belonging to GH53. It also revealed 15 zinc ions bound to the protein, one of which is located in the active site, where it is coordinated by residues Glu136 and Glu246 which comprise the catalytic machinery. The majority of the zinc ions are located on the surface of the enzyme, in some cases with side chains from two different molecules as ligands, thus explaining why the presence of zinc ions was essential for crystallization.
authors.affiliation: Department of Chemistry, University of Copenhagen, Universitetsparken 5, DK-2100 Copenhagen, Denmark.
authors.forename: Harm
authors.initials: H
authors.lastname: Otten
authors.forename: Malwina
authors.initials: M
authors.lastname: Michalak
authors.forename: Jørn Dalgaard
authors.initials: JD
authors.lastname: Mikkelsen
authors.forename: Sine
authors.initials: S
authors.lastname: Larsen
mesh_terms: D000595:Amino Acid Sequence; D001665:Binding Sites / Q000502:physiology; D018360:Crystallography, X-Ray; D032842:Emericella / Q000201:enzymology / Q000235:genetics; D005656:Fungal Proteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D006026:Glycoside Hydrolases / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D008969:Molecular Sequence Data; D017433:Protein Structure, Secondary; D017434:Protein Structure, Tertiary; D014961:X-Ray Diffraction; D015032:Zinc / Q000737:chemistry / Q000378:metabolism*
chemical_list: D005656:Fungal Proteins; D006026:Glycoside Hydrolases; C094192:arabinogalactan endo-1,4-beta-galactosidase; D015032:Zinc
keywords: crystallization; family GH53; galactanases; glycosyl hydrolases; zinc binding
references.citation: Appl Microbiol Biotechnol. 2011 May;90(3):873-84
references.pmid: 21253720
references.citation: Acta Crystallogr D Biol Crystallogr. 2012 Sep;68(Pt 9):1253-8
references.pmid: 22948927
references.citation: J Synchrotron Radiat. 2009 Nov;16(Pt 6):872-9
references.pmid: 19844027
references.citation: Acta Crystallogr D Biol Crystallogr. 2010 Jan;66(Pt 1):12-21
references.pmid: 20057044
references.citation: Nucleic Acids Res. 2009 Jan;37(Database issue):D233-8
references.pmid: 18838391
references.citation: J Mol Biol. 2004 Jul 30;341(1):107-17
references.pmid: 15312766
references.citation: Acta Crystallogr D Biol Crystallogr. 2006 Jan;62(Pt 1):116-24
references.pmid: 16369101
references.citation: FEBS Lett. 1995 Apr 10;362(3):281-5
references.pmid: 7729513
references.citation: Biochemistry. 2002 Dec 24;41(51):15135-43
references.pmid: 12484750
references.citation: Science. 1997 Sep 12;277(5332):1662-6
references.pmid: 9287219
references.citation: Acta Crystallogr D Biol Crystallogr. 2004 May;60(Pt 5):849-59
references.pmid: 15103130
references.citation: Proc Natl Acad Sci U S A. 1995 Jul 18;92(15):7090-4
references.pmid: 7624375
references.citation: Enzyme Microb Technol. 2012 Feb 10;50(2):121-9
references.pmid: 22226198
references.citation: Enzyme Microb Technol. 2011 Aug 10;49(3):289-97
references.pmid: 22112514
references.citation: Biotechnol Bioeng. 2007 Oct 15;98(3):711-8
references.pmid: 17461426
references.citation: Acta Crystallogr D Biol Crystallogr. 2010 Feb;66(Pt 2):133-44
references.pmid: 20124693
references.citation: Protein Sci. 2003 Jun;12(6):1195-204
references.pmid: 12761390
references.citation: J Biotechnol. 2013 Mar 10;164(1):75-81
references.pmid: 23313889
references.citation: Proteins. 2009 Jun;75(4):977-89
references.pmid: 19089956
references.citation: Proc Natl Acad Sci U S A. 2006 Jul 25;103(30):11417-22
references.pmid: 16844780
references.citation: J Synchrotron Radiat. 2010 Jan;17(1):107-18
references.pmid: 20029119
references.citation: Enzyme Microb Technol. 2011 Jul 10;49(2):160-6
references.pmid: 22112403