title: The crystal structure of galactitol-1-phosphate 5-dehydrogenase from Escherichia coli K12 provides insights into its anomalous behavior on IMAC processes.
abstract: Endogenous galactitol-1-phosphate 5-dehydrogenase (GPDH) (EC 1.1.1.251) from Escherichia coli spontaneously interacts with Ni(2+)-NTA matrices becoming a potential contaminant for recombinant, target His-tagged proteins. Purified recombinant, untagged GPDH (rGPDH) converted galactitol into tagatose, and d-tagatose-6-phosphate into galactitol-1-phosphate, in a Zn(2+)- and NAD(H)-dependent manner and readily crystallized what has permitted to solve its crystal structure. In contrast, N-terminally His-tagged GPDH was marginally stable and readily aggregated. The structure of rGPDH revealed metal-binding sites characteristic from the medium-chain dehydrogenase/reductase protein superfamily which may explain its ability to interact with immobilized metals. The structure also provides clues on the harmful effects of the N-terminal His-tag.
authors.affiliation: Laboratorio de Biotecnología Bacteriana, Instituto Ciencia y Tecnología de Alimentos y Nutrición (ICTAN), CSIC, Madrid, Spain.
authors.forename: María
authors.initials: M
authors.lastname: Esteban-Torres
authors.forename: Yanaisis
authors.initials: Y
authors.lastname: Alvarez
authors.forename: Iván
authors.initials: I
authors.lastname: Acebrón
authors.forename: Blanca
authors.initials: B
authors.lastname: de las Rivas
authors.forename: Rosario
authors.initials: R
authors.lastname: Muñoz
authors.forename: Gert-Wieland
authors.initials: GW
authors.lastname: Kohring
authors.forename: Ana María
authors.initials: AM
authors.lastname: Roa
authors.forename: Mónica
authors.initials: M
authors.lastname: Sobrino
authors.forename: José M
authors.initials: JM
authors.lastname: Mancheño
mesh_terms: D000595:Amino Acid Sequence; D001483:Base Sequence; D001665:Binding Sites; D002846:Chromatography, Affinity; D018360:Crystallography, X-Ray; D004269:DNA, Bacterial / Q000235:genetics; D004795:Enzyme Stability; D048168:Escherichia coli K12 / Q000201:enzymology* / Q000235:genetics; D029968:Escherichia coli Proteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D005798:Genes, Bacterial; D008670:Metals / Q000378:metabolism; D008958:Models, Molecular; D008969:Molecular Sequence Data; D055503:Protein Multimerization; D011994:Recombinant Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D017386:Sequence Homology, Amino Acid; D013401:Sugar Alcohol Dehydrogenases / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism
chemical_list: D004269:DNA, Bacterial; D029968:Escherichia coli Proteins; D008670:Metals; D011994:Recombinant Proteins; D013401:Sugar Alcohol Dehydrogenases; C031269:L-galactitol-1-phosphate dehydrogenase