The crystal structure of galactitol-1-phosphate 5-dehydrogenase from Escherichia coli K12 provides insights into its anomalous behavior on IMAC processes

作者：Esteban Torres Maria; Alvarez Yanaisis; Acebron Ivan; de las Rivas Blanca; Munoz Rosario; Kohring Gert Wieland; Maria Roa Ana; Sobrino Monica; Mancheno Jose M^*

来源：FEBS Letters, 2012, 586(19): 3127-3133.

DOI：10.1016/j.febslet.2012.07.073

• Crystal structure
• Dehydrogenase
• D-Tagatose-6-phosphate
• Galactitol
• Metal binding-site
• Zn-metalloenzyme

Endogenous galactitol-1-phosphate 5-dehydrogenase (GPDH) (EC 1.1.1.251) from Escherichia coli spontaneously interacts with Ni2+-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 Zn2+- 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. %26lt;br%26gt;Structured summary of protein interactions: %26lt;br%26gt;GPDH and GPDH bind by molecular sieving (View interaction) %26lt;br%26gt;GPDH and GPDH bind by x-ray crystallography (Vie

• 出版日期2012-9-21