The Salk Institute for Biological Studies

Toudai Tlo, Ltd.

Stephane Richard

Joseph Noel

Tomohisa Kuzuyama

Marianne Bowman

Elise Larsen

Florence Pojer

CHI like fatty acid binding proteins and genes, recombinant cells and organisms, methods of metabolic pathway engineering to improve lipid production in cells, Crystal structures of CHI like fatty acid binding proteins, methods of engineering CHI like fatty acid binding proteins and systems thereof are provided.

Metabolic engineering of lipid metabolism by improving fatty acid binding and transport

In accordance with the present invention, a novel aromatic prenyltransferase, Orf2 fromsp. strain CL190, involved in naphterpin biosynthesis has been identified and the structure thereof elucidated. This prenyltransferase catalyzes the formation of a C&#x2014;C bond between a prenyl group and a compound containing an aromatic nucleus, and also displays C&#x2014;O bond formation activity. Numerous crystallographic structures of the prenyltransferase have been solved and refined, e.g., (1) prenyltransferase complexed with a buffer molecule (TAPS), (2) prenyltransferase as a binary complex with geranyl diphosphate (GPP) and Mg, and prenyltransferase as ternary complexes with a non-hydrolyzable substrate analogue, geranyl S-thiolodiphosphate (GSPP) and either (3) 1,6-dihydroxynaphthalene (1,6-DHN), or (4) flaviolin (i.e., 2,5,7-trihydroxy-1,4-naphthoquinone, which is the oxidized product of 1,3,6,8-tetrahydroxynaphthalene (THN)). These structures have been solved and refined to 1.5 &#x212b;, 2.25 &#x212b;, 1.95 &#x212b; and 2.02 &#x212b;, respectively. This first structure of an aromatic prenyltransferase displays an unexpected and non-canonical (&#x3b2;/&#x3b1;)-barrel architecture. The complexes with both aromatic substrates and prenyl containing substrates and analogs delineate the active site and are consistent with a proposed electrophilic mechanism of prenyl group transfer. These structures also provide a mechanistic basis for understanding prenyl chain length determination and aromatic co-substrate recognition in this structurally unique family of aromatic prenyltransferases. This structural information is useful for predicting the aromatic prenyltransferase activity of proteins.

Aromatic prenyltransferases, nucleic acids encoding same and uses therefor

The present invention provides the structure of the enzyme 4-diphosphocytidyl-2-C-methylerythritol (CDP-ME) synthase, a member of the cytidyltransferase family of enzymes from. CDP-ME is a critical intermediate in the mevalonate-independent pathway for isoprenoid biosynthesis in a number of prokaryotic organisms, in algae, in the plastids of plants, and in the malaria parasite. Since vertebrates synthesize isoprenoid precursors using a mevalonate pathway, CDP-ME synthase and other enzymes of the mevalonate-independent pathway for isoprenoid production represent attractive targets for the structure-based design of selective antibacterial, herbicidal, and antimalarial drugs. Accordingly, the present invention provides methods for screening for compounds that inhibit enzymes of the mevalonate-independent pathway and pharmaceutical compositions and antibacterial formulations thereof. Further provided are methods of inhibiting the enzymes of the pathway and bacterial terpenoid synthesis and methods for treating a subject suffering from a bacterial infection.

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Del Mar, CA, United States of America

Stephane Richard