Paper Citing NAMD - Abstract
Joseph, Dmitri D. A.; Jiao, Wanting; Kessans, Sarah A.; Parker, Emily J.
Substrate-mediated control of the conformation of an ancillary domain delivers a competent catalytic site for N-acetylneuraminic acid synthase
PROTEINS-STRUCTURE FUNCTION AND BIOINFORMATICS, 82:2054-2066, SEP 2014
N-Acetylneuratiiinic acid (NANA) is the most common naturally occurring sialic acid and plays a key role in the pathogenesis of a select number of nettroinvasive bacteria such as Neisseria meningitidis. NANA is synthesized in prokaryotes via a condensation reaction between phosphoentalpyruvate and N-acetylrnannosatnirie. This reaction is catalyzed by a domain swapped, homodirneric enzyme, N-acetyltieurarriiiiic acid sluithase (NANIAS). NANIAS comprises two distinct domains an Nterminal catalytic (beta/alpha)8 barrel linked to a C-terminal antifreeze protein-like (AFPL) domain. We have investigated the role of the AFPL domain by characterizing a truncated variant of NrtieNANAS, which was discovered to be soluble yet inactive. Analytical ttltracentrifugation and analytical size exclusion were used to probe the quaternary state of the NtneNANAS truncation, and revealed that loss of the AFPL domain destabilizes the dimeric form of the enzyme. The results from this study thereby demonstrate that the AFPL domain plays a critical role for both the catalytic function and quaternary structure stability of NANAS. Small angle X-ray scattering, molecular dynamics simulations, and amino acid substitutions expose a complex hydrogen-bonding relay, which links the roles of the catalytic and AFPL domains across subunit boundaries.
