3QPY

Crystal structure of a mutant (K57A) of 3-deoxy-D-manno-octulosonate 8-phosphate synthase (KDO8PS) from Neisseria meningitidis

Summary for 3QPY

• Entry DOI: 10.2210/pdb3qpy/pdb
• Related: 3QPZ 3QQ0 3QQ1
• Descriptor: 2-dehydro-3-deoxyphosphooctonate aldolase, CHLORIDE ION, SODIUM ION, ... (5 entities in total)
• Functional Keywords: manno-octulosonate, synthase, lipopolysaccharide, kdop, kdo8 kdops, kdo8ps, tim barrel, biosynthesis, transferase, lyase, lipopolysaccharide biosynthesis
• Biological source: Neisseria meningitidis
• Cellular location: Cytoplasm (By similarity): Q9JZ55
• Total number of polymer chains: 4
• Total formula weight: 122066.34

Authors

Allison, T.M.,Jameson, G.B.,Parker, E.J. (deposition date: 2011-02-14, release date: 2011-04-13, Last modification date: 2023-11-01)

Primary citation

Allison, T.M.,Hutton, R.D.,Cochrane, F.C.,Yeoman, J.A.,Jameson, G.B.,Parker, E.J.
Targeting the role of a key conserved motif for substrate selection and catalysis by 3-deoxy-D-manno-octulosonate 8-phosphate synthase
Biochemistry, 50:3686-3695, 2011

PubMed Abstract: 3-Deoxy-D-manno-octulosonate 8-phosphate synthase (KDO8PS) catalyzes the reaction between three-carbon phosphoenolpyruvate (PEP) and five-carbon d-arabinose 5-phosphate (A5P), generating KDO8P, a key intermediate in the biosynthetic pathway to 3-deoxy-D-manno-octulosonate, a component of the lipopolysaccharide of the Gram-negative bacterial cell wall. Both metal-dependent and metal-independent forms of KDO8PS have been characterized. KDO8PS is evolutionarily and mechanistically related to the first enzyme of the shikimate pathway, the obligately divalent metal ion-dependent 3-deoxy-D-arabino-heptulosonate 7-phosphate synthase (DAH7PS) that couples PEP and four-carbon D-erythrose 4-phosphate (E4P) to give DAH7P. In KDO8PS, an absolutely conserved KANRS motif forms part of the A5P binding site, whereas in DAH7PS, an absolutely conserved KPR(S/T) motif accommodates E4P. Here, we have characterized four mutants of this motif (AANRS, KAARS, KARS, and KPRS) in metal-dependent KDO8PS from Acidithiobacillus ferrooxidans and metal-independent KDO8PS from Neisseria meningitidis to test the roles of the universal Lys and the Ala-Asn portion of the KANRS motif. The X-ray structures, determined for the N. meningitidis KDO8PS mutants, indicated no gross structural penalty resulting from mutation, but the subtle changes observed in the active sites of these mutant proteins correlated with their altered catalytic function. (1) The AANRS mutations destroyed catalytic activity. (2) The KAARS mutations lowered substrate selectivity, as well as activity. (3) Replacing KANRS with KARS or KPRS destroyed KDO8PS activity but did not produce a functional DAH7PS. Thus, Lys is critical to catalysis, and other changes are necessary to switch substrate specificity for both the metal-independent and metal-dependent forms of these enzymes.

PubMed: 21438567
DOI: 10.1021/bi200251f

Experimental method

X-RAY DIFFRACTION (1.95 Å)