Structural and Kinetic Characterization of the LPS Biosynthetic Enzyme D-alpha,beta-D-heptose-1,7-bisphosphate Phosphatase (GmhB) from Escherichia coli

Journal Article · Fri Jan 01 04:00:00 EST 2010 · Biochemistry

DOI:https://doi.org/10.1021/bi901780j· OSTI ID:1019856

Taylor, P; Sugiman-Marangos, S; Zhang, K; Valvano, M; Wright, G; Junop, M

Lipopolysaccharide is a major component of the outer membrane of Gram-negative bacteria and provides a permeability barrier to many commonly used antibiotics. ADP-heptose residues are an integral part of the LPS inner core, and mutants deficient in heptose biosynthesis demonstrate increased membrane permeability. The heptose biosynthesis pathway involves phosphorylation and dephosphorylation steps not found in other pathways for the synthesis of nucleotide sugar precursors. Consequently, the heptose biosynthetic pathway has been marked as a novel target for antibiotic adjuvants, which are compounds that facilitate and potentiate antibiotic activity. D-{alpha},{beta}-D-Heptose-1,7-bisphosphate phosphatase (GmhB) catalyzes the third essential step of LPS heptose biosynthesis. This study describes the first crystal structure of GmhB and enzymatic analysis of the protein. Structure-guided mutations followed by steady state kinetic analysis, together with established precedent for HAD phosphatases, suggest that GmhB functions through a phosphoaspartate intermediate. This study provides insight into the structure-function relationship of GmhB, a new target for combatting Gram-negative bacterial infection.

• Research Organization: Brookhaven National Laboratory (BNL) National Synchrotron Light Source
• Sponsoring Organization: DOE - OFFICE OF SCIENCE
• DOE Contract Number: AC02-98CH10886
• OSTI ID: 1019856
• Report Number(s): BNL--95702-2011-JA
• Journal Information: Biochemistry, Journal Name: Biochemistry Journal Issue: 5 Vol. 49; ISSN 0006-2960
• Country of Publication: United States
• Language: English

Related Subjects

59 BASIC BIOLOGICAL SCIENCES
60 APPLIED LIFE SCIENCES
ANTIBIOTICS
BACTERIA
BIOSYNTHESIS
CRYSTAL STRUCTURE
ENZYMES
ESCHERICHIA COLI
KINETICS
LIPOPOLYSACCHARIDES
MEMBRANES
MUTANTS
MUTATIONS
NUCLEOTIDES
PERMEABILITY
PHOSPHATASES
PHOSPHORYLATION
RESIDUES
SACCHAROSE
SYNTHESIS
TARGETS
national synchrotron light source