SOLUTION STRUCTURE OF THE ACTINORHODIN POLYKETIDE SYNTHASE ACYL CARRIER PROTEIN FROM STREPTOMYCES-COELICOLOR A3(2)

The solution structure of the actinorhodin acyl carrier protein ( net apo-ACP) from the polyketide synthase (PKS) of Streptomyces coelicolor A3(2) has been determined using H-1 NMR spectroscopy, representing th e first polyketide synthase component for which detailed structural in formation has been obtained. Twenty-four structures were generated by simulated annealing, employing 699 distance restraints and 94 dihedral angle restraints. The structure is composed, principally, of three ma jor helices (1, 2, and 4), a shorter helix (3) and a large loon region separating helices 1 and 2. The structure is well-defined, except for a portion of the loop region (residues 18-29), the N-terminus (1-4), and a short stretch (57-61) in the loop connecting helices 2 and 3. Th e RMS distribution of the 24 structures about the average structure is 1.47 Angstrom for backbone atoms, 1.84 Angstrom for all heavy atoms ( residues 5-86), and 1.01 Angstrom for backbone atoms over the helical regions (5-18, 41-86). The tertiary fold of act apo-ACP shows a strong structural homology with Escherichia coli fatty acid synthase (FAS) A CP, though some structural differences exist. First, there is no evide nce that act apo-ACP is conformationally averaged between two or more states as observed in E. coli FAS ACP. Second, act apo-ACP shows a dis ordered N-terminus (residues 1-4) and a longer flexible loop (19-41 wi th 19-29 disordered) as opposed to E. coli FAS ACP where the N-termina l helix starts at residue 3 and the loop region is three amino acids s horter (16-35). Most importantly, however, although the act apo-ACP st ructure contains a hydrophobic core, there are in addition a number of buried hydrophilic groups, principally Arg72 and Asn79, both of which are 100% conserved in the PKS ACPs and not the FAS ACPs and may there fore play a role in stabilizing the growing polyketide chain. The stru cture-function relationship of act ACP is discussed in the light of th ese structural data and recent genetic advances in the field.