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Structural and biochemical characterization of the class II fructose‐1,6‐bisphosphatase from Francisella tularensis
Author(s) -
Selezneva Anna I.,
Gutka Hiten J.,
Wolf Nina M.,
Qurratulain Fnu,
Movahedzadeh Farahnaz,
Abad-Zapatero Celerino
Publication year - 2020
Publication title -
acta crystallographica section f
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.572
H-Index - 37
ISSN - 2053-230X
DOI - 10.1107/s2053230x20013370
Subject(s) - homotetramer , active site , groes , crystallography , chemistry , allosteric regulation , stereochemistry , biology , biochemistry , escherichia coli , enzyme , groel , protein subunit , gene
The crystal structure of the class II fructose‐1,6‐bisphosphatase (FBPaseII) from the important pathogen Francisella tularensis is presented at 2.4 Å resolution. Its structural and functional relationships to the closely related phosphatases from Mycobacterium tuberculosis ( Mt FBPaseII) and Escherichia coli ( Ec FBPaseII) and to the dual phosphatase from Synechocystis strain 6803 are discussed. FBPaseII from F. tularensis ( Ft FBPaseII) was crystallized in a monoclinic crystal form (space group P 2 1 , unit‐cell parameters a = 76.30, b = 100.17, c = 92.02 Å, β = 90.003°) with four chains in the asymmetric unit. Chain A had two coordinated Mg 2+ ions in its active center, which is distinct from previous findings, and is presumably deactivated by their presence. The structure revealed an approximate 222 ( D 2 ) symmetry homotetramer analogous to that previously described for Mt FBPaseII, which is formed by a crystallographic dyad and which differs from the exact tetramer found in Ec FBPaseII at a 222 symmetry site in the crystal. Instead, the approximate homotetramer is very similar to that found in the dual phosphatase from Synechocystis , even though no allosteric effector was found in Ft FBPase. The amino‐acid sequence and folding of the active site of Ft FBPaseII result in structural characteristics that are more similar to those of the previously published Ec FBPaseII than to those of Mt FBPaseII. The kinetic parameters of native Ft FBPaseII were found to be in agreement with published studies. Kinetic analyses of the Thr89Ser and Thr89Ala mutations in the active site of the enzyme are consistent with the previously proposed mechanism for other class II bisphosphatases. The Thr89Ala variant enzyme was inactive but the Thr89Ser variant was partially active, with an approximately fourfold lower K m and V max than the native enzyme. The structural and functional insights derived from the structure of Ft FBPaseII will provide valuable information for the design of specific inhibitors.