title: Structure of glycerol dehydrogenase from Serratia.
abstract: The 1.90 Å resolution X-ray crystal structure of glycerol dehydrogenase derived from contaminating bacteria present during routine Escherichia coli protein expression is presented. This off-target enzyme showed intrinsic affinity for Ni(2+)-Sepharose, migrated at the expected molecular mass for the target protein during gel filtration and was crystallized before it was realised that contamination had occurred. In this study, it is shown that liquid chromatography coupled to tandem mass spectrometry (LC-MS/MS) can efficiently identify the protein composition of crystals in a crystallization experiment as part of a structure-determination pipeline for an unknown protein. The high-resolution X-ray data enabled sequencing directly from the electron-density maps, allowing the source of contamination to be placed within the Serratia genus. Incorporating additional protein-identity checks, such as tandem LC-MS/MS, earlier in the protein expression, purification and crystallization workflow may have prevented the unintentional structure determination of this metabolic enzyme, which represents the first enterobacterial glycerol dehydrogenase reported to date.
authors.affiliation: Department of Biochemistry, Emory University School of Medicine, 1510 Clifton Road RRC G235, Atlanta, GA 30033, USA.
authors.forename: Paul
authors.initials: P
authors.lastname: Musille
authors.affiliation: Department of Biochemistry, Emory University School of Medicine, 1510 Clifton Road RRC G235, Atlanta, GA 30033, USA.
authors.forename: Eric
authors.initials: E
authors.lastname: Ortlund
mesh_terms: D002850:Chromatography, Gel; D003460:Crystallization; D018360:Crystallography, X-Ray; D008970:Molecular Weight; D011487:Protein Conformation; D012705:Serratia / Q000201:enzymology*; D013401:Sugar Alcohol Dehydrogenases / Q000737:chemistry*; D053719:Tandem Mass Spectrometry
chemical_list: D013401:Sugar Alcohol Dehydrogenases; C020672:glycerol dehydrogenase
keywords: Serratia; glycerol dehydrogenase