AN EVALUATION OF MOLECULAR-MODELS OF THE CYTOCHROME-P450 STREPTOMYCES-GRISEOLUS ENZYMES P450SU1 AND P450SU2

P450SU1 and P450SU2 are herbicide-inducible bacterial cytochrome P450 enzymes from Streptomyces griseolus. They have two of the highest sequ ence identities to camphor hydroxylase (P450cam from Pseudomonas putid a), the cytochrome P450 with the first known crystal structure. We hav e built several models of these two proteins to investigate the variab ility in the structures that can occur from using different modeling p rotocols. We looked at variability due to alignment methods, backbone loop conformations and refinement methods. We have constructed two mod els for each protein using two alignment algorithms, and then an addit ional model using an identical alignment but different loop conformati ons for both buried and surface loops. The alignments used to build th e models were created using the Needleman-Wunsch method, adapted for m ultiple sequences, and a manual method that utilized both a dot-matrix search matrix and the Needleman-Wunsch method. After constructing the initial models, several energy minimization methods were used to expl ore the variability in the final models caused by the choice of minimi zation techniques. Features of cytochrome P450cam and the cytochrome P 450 superfamily, such as the ferredoxin binding site, the heme binding site and the substrate binding site were used to evaluate the validit y of the models. Although the final structures were very similar betwe en the models with different alignments, active-site residues were fou nd to be dependent on the conformations of buried loops and early stag es of energy minimization. We show which regions of the active site ar e the most dependent on the particular methods used, and which parts o f the structures seem to be independent of the methods.