z-logo
Premium
Crystal structure of a metal‐dependent phosphoesterase (YP_910028.1) from Bifidobacterium adolescentis : Computational prediction and experimental validation of phosphoesterase activity
Author(s) -
Han Gye Won,
Ko Jaeju,
Farr Carol L.,
Deller Marc C.,
Xu Qingping,
Chiu HsiuJu,
Miller Mitchell D.,
Sefcikova Jana,
Somarowthu Srinivas,
Beuning Penny J.,
Elsliger MarcAndré,
Deacon Ashley M.,
Godzik Adam,
Lesley Scott A.,
Wilson Ian A.,
Ondrechen Mary Jo
Publication year - 2011
Publication title -
proteins: structure, function, and bioinformatics
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.699
H-Index - 191
eISSN - 1097-0134
pISSN - 0887-3585
DOI - 10.1002/prot.23035
Subject(s) - polymerase , dna polymerase , dna , primer (cosmetics) , biology , taq polymerase , computational biology , chemistry , biochemistry , thermus aquaticus , organic chemistry
The crystal structures of an unliganded and adenosine 5′‐monophosphate (AMP) bound, metal‐dependent phosphoesterase (YP_910028.1) from Bifidobacterium adolescentis are reported at 2.4 and 1.94 Å, respectively. Functional characterization of this enzyme was guided by computational analysis and then confirmed by experiment. The structure consists of a polymerase and histidinol phosphatase (PHP, Pfam: PF02811) domain with a second domain (residues 105‐178) inserted in the middle of the PHP sequence. The insert domain functions in binding AMP, but the precise function and substrate specificity of this domain are unknown. Initial bioinformatics analyses yielded multiple potential functional leads, with most of them suggesting DNA polymerase or DNA replication activity. Phylogenetic analysis indicated a potential DNA polymerase function that was somewhat supported by global structural comparisons identifying the closest structural match to the alpha subunit of DNA polymerase III. However, several other functional predictions, including phosphoesterase, could not be excluded. Theoretical microscopic anomalous titration curve shapes, a computational method for the prediction of active sites from protein 3D structures, identified potential reactive residues in YP_910028.1. Further analysis of the predicted active site and local comparison with its closest structure matches strongly suggested phosphoesterase activity, which was confirmed experimentally. Primer extension assays on both normal and mismatched DNA show neither extension nor degradation and provide evidence that YP_910028.1 has neither DNA polymerase activity nor DNA‐proofreading activity. These results suggest that many of the sequence neighbors previously annotated as having DNA polymerase activity may actually be misannotated. Proteins 2011. © 2011 Wiley‐Liss, Inc.

This content is not available in your region!

Continue researching here.

Having issues? You can contact us here