This subproject is one of many research subprojects utilizing theresources provided by a Center grant funded by NIH/NCRR. The subproject andinvestigator (PI) may have received primary funding from another NIH source,and thus could be represented in other CRISP entries. The institution listed isfor the Center, which is not necessarily the institution for the investigator.In E. coli and other Gram-negative bacteria, KDO (3-deoxy-D-manno-octulosonate), a component of the lipopolysaccharide inner core, is essential for cell viability. KDO 8-P phosphatase (YrbI), which catalyzes the hydrolysis of KDO 8-P to KDO and inorganic phosphate, is the last enzyme in the KDO biosynthetic pathway for which the gene has recently been identified. The mechanism of its divalent metal-dependent phosphatase activity has not yet been elucidated. We have obtained crystals of YrbI, which diffract well (some to about 2 A) using our in-house Rigaku rotating anode generator. We have crystals of YrbI with different divalent metals, those soaked or cocrystallized with KDO substrate (at conditions where reaction is either slowed down or with divalent metals (calcium), which do not support catalysis). A synchrotron source is absolutely critical in providing high resolution data, which will allow us to see details in the active site for different protein-divalent metal-substrate complexes.
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