The crystal structure of chorismate lyase shows a new fold and a tightly retained product

The enzyme chorismate lyase (CL) catalyzes the removal of pyruvate from cho rismate to produce 4-hydroxy benzoate (4HB) for the ubiquinone pathway. In Escherichia coli, CL is monomeric, with 164 residues. We have determined th e structure of the CL product complex by crystallographic heavy-atom method s and report the structure at 1.4-Angstrom resolution for a fully active do uble Cys-to-Ser mutant and at 2.0-Angstrom resolution for the wild-type. Th e fold involves a 6-stranded antiparallel beta -sheet with no spanning heli ces and novel connectivity. The product is bound internally, adjacent to th e sheet, with its polar groups coordinated by two main-chain amides and by the buried sidechains of Arg 76 and Glu 155. The 4HB is completely sequeste red from solvent in a largely hydrophobic environment behind two helix-turn -helix loops. The extensive product binding that is observed is consistent with biochemical measurements of slow product release and 10-fold stronger binding of product than substrate. Substrate binding and kinetically rate-l imiting product release apparently require the rearrangement of these activ e-site-covering loops. Implications for the biological function of the high product binding are considered in light of the unique cellular role of 4HB , which is produced by cytoplasmic CL but is used by the membrane-bound enz yme 4HB octaprenyltransferase. (C) 2001 Wiley-Liss, Inc.*