Book:Structural Proteomics and its Impact on the Life Sciences:6
Chapter 6: The Contribution of Structural Proteomics to Understanding the Function of Hypothetical ProteinsMichael D. Suits, Allan Matte, Zongchao Jia and Miroslaw Cygler [1] Molecular Tour A quinate/shikimate dehydrogenase from Escherichia coli - YdiB YdiB, an E. coli protein, was annotated as a putative shikimate dehydrogenase (1o9b [3], monomer A is in magenta, monomer B is in cyan), as it possessed 28% sequence identity to E. coli AroE, a characterized shikimate dehydrogenase[4]. The structure of E. coli YdiB revealed interesting similarities and differences to E. coli AroE (1nyt). YdiB is in magenta, AroE is in yellow, carbon atoms of NAD and NADP are in green. Click here to see animation of this scene. Both enzymes share similar folds, contain signature motifs indicative of dinucleotide (NAD+, NADP+) binding, and while AroE is a monomer, the crystal structure of E. coli YdiB revealed a dimer [3]. The mode of dimerization of YdiB is unusual, as it involves a relatively small surface of each monomer, but is consistent with the molecular weight determined by size exclusion chromatography. Both enzymes consist of two α/β domains, with the C-terminal domain adopting a canonical Rossmann fold topology, consistent with dinucleotide binding. Escherichia coli Proteins Associated With Heme Metabolism At the initiation of our investigation ChuS, from Shigella dysenteriae (1u9t), was known only as a protein up-regulated under low iron conditions, responsible for the prevention of heme toxicity. It showed low sequence homology to know 3D structures and thus there was no clear function for this protein. The structure of ChuS revealed a central core of two, 9-stranded anti-parallel β-sheets, each flanked at its N-terminus by one pair of parallel α-helices and at its C-terminus by a set of three α-helices in an helix-loop-helix-loop-helix configuration. N- and C-terminal domains are colored red and blue, respectively. The structure clearly indicated that ChuS arose by gene duplication, as can be seen by superimposing the N and C terminal domains on each other resulting in an RMS of of 2.1 Å. Based on the ChuS site directed mutation studies were carried out that indicated that demonstrate in the ChuS-heme complex (2hq2, N- and C-terminal domains are colored indigo and green, respectively) that His193 is responsible for axial ligation of heme, and that Arg100 further stabilizes the heme group from the medial side of the protein via two water molecules. Carbon atoms of heme are in yellow, Arg100 and His193 are in magenta, and water molecules are in red. It is also interesting to note that the dimer structure of ChuX (2ovi) displays a surprising degree of similarity to the monomer structures of two other heme utilization operon proteins, E. coli ChuS and Y. enterocolitica HemS, both of which have an internal domain repeat. The conserved His65 and His98 are in yellow and red, respectively.
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