In bacteria, the shikimate pathway-derived intermediate chorismate is an important branch point for the synthesis of a number of primary and secondary metabolites. Examples include the aromatic amino acids, folate, coenzymes Q and the antibiotics chloramphenicol and candicidin, to name a few. There are five known chorismate-utilizing enzymes that catalyze reactions in pathways that branch off from chorismate. Two of these are anthranilate synthase with its associated aminodeoxyisochorismate (ADIC) synthase activity, and p-aminobenzoic acid (PABA) synthase with its associated aminodeoxychorismate (ADC) activity. Most anthranilate syntheses involved in tryptophan biosynthesis have distinct TrpE and TrpG subunits;ADC synthase contains PabA and PabB subunits. The work proposed here is directed toward the characterization or engineering of fused chorismate-utilizing enzymes. One such enzyme is from Streptomyces venezuelae (Sv), a chloramphenicol-producing Gram positive bacterium. A second is from Streptomyces griseus, producer of the antifungal antibiotic candicidin (and streptomycin). A third originates in Escherichia coli. There are three major aims or goals: (1) the detailed characterization of the structure and function of the fused SvTrpEG, which, contrary to initial expectations, has been found to be an anthranilate synthase as opposed to an ADIC synthase;(2) the cloning, expression and preliminary characterization of PabAB from S. griseus;and (3) the engineering of a fused E. coli PabAB enzyme. Achievement of the first goal will provide a deeper understanding of what structurally and functionally sets TrpEG apart from the homologous phenazine biosynthetic enzyme PhzE. Achievement of goal 2 will further an understanding of how PabA and PabB subunits of the metabolically important enzyme PABA synthase associate and work together to produce ADC, from which PABA is formed through the catalytic action of a third enzyme, ADC lyase or PabC. It could also aid in the metabolic engineering of the candicidin biosynthetic pathway in order to increase yield of antibiotic. Finally, achievement of the third goal may solve a problem that has plagued the study of the interaction between the PabA and PabB subunits of E. coli PABA synthase: their weak interaction when separate.

National Institute of Health (NIH)
National Institute of General Medical Sciences (NIGMS)
Research Continuance Award (SC3)
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Special Emphasis Panel (ZGM1-MBRS-X (CH))
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Okita, Richard T
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Howard University
Schools of Medicine
United States
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Byrnes, W Malcolm (2015) E. E. Just and Creativity in Science. The Importance of Diversity. J Afr Am Stud (New Brunsw) 19:264-278
Byrnes, W Malcolm; Newman, Stuart A (2014) Ernest Everett Just: Egg and embryo as excitable systems. J Exp Zool B Mol Dev Evol 322:191-201
Ashenafi, Meseret; Ammosova, Tatiana; Nekhai, Sergei et al. (2014) Purification and characterization of aminoglycoside phosphotransferase APH(6)-Id, a streptomycin-inactivating enzyme. Mol Cell Biochem 387:207-16
Debebe, Zufan; Nekhai, Sergei; Ashenafi, Meseret et al. (2012) Development of a sensitive HPLC method to measure in vitro permeability of E- and Z-isomeric forms of thiosemicarbazones in Caco-2 monolayers. J Chromatogr B Analyt Technol Biomed Life Sci 906:25-32
Byrnes, W Malcolm (2009) Ernest Everett Just, Johannes Holtfreter, and the origin of certain concepts in embryo morphogenesis. Mol Reprod Dev 76:912-21