A proposal for the catalytic mechanism in phospholipase C based on interaction energy and distance geometry calculations.
The non-specific phospholipase C from Bacillus cereus preferentially hydrolyses phosphatidylcholine but is also active against phosphatidylserine, phosphatidylethanolamine and at a much lower level, sphingomyelin. A minimal substrate model containing all required structural and configurational elements of a high affinity substrate was docked into the active site. The enzyme-substrate attachment points were from molecular interaction energy calculations using the program GRID and from a previous phosphate inhibitor complex structure. Available conformational space for the substrate was sampled by distance geometry calculations using the program DGEOM. This investigation clearly identifies the attacking nucleophile, a catalytically favourable orientation of the phosphate group in its tetra-, as well as its penta-, coordinated state and a crucial stabilizing environment for the alkoxide intermediate. Based on this information a complete catalytic cycle is proposed.
Department of Medicinal Biochemistry, University of Göteborg, Sweden.
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Sundell
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Hansen
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Hough
D001409:Bacillus cereus / Q000201:enzymology*; D001665:Binding Sites; D002384:Catalysis; D018360:Crystallography, X-Ray; D008432:Mathematical Computing; D008954:Models, Biological; D008956:Models, Chemical; D008968:Molecular Conformation; D010712:Phosphatidic Acids / Q000378:metabolism; D013109:Sphingomyelins / Q000378:metabolism; D013379:Substrate Specificity; D010738:Type C Phospholipases / Q000737:chemistry* / Q000378:metabolism*
D010712:Phosphatidic Acids; D013109:Sphingomyelins; D010738:Type C Phospholipases