E0002.html

Time-resolved studies can be one of two types; an experimental time sequence or a computer interpolation between two end state static crystal structures. Closely allied to the first type is the temperature-resolved study. Time-resolved experiments can be based on diffusion of an enzyme substrate or pH jump induced or light induced. The desire to see a molecule in action is the vision of all these types of experiment. The molecular structure may well be constrained by the crystal lattice interactions. Further insights can be gained from molecular dynamics studies into the pathway of structures. These topics will be covered in the sessions as a whole at ACA 2003. From my Lab in Manchester two case studies will be described in detail. Firstly the determination of several structures of the active form of the enzyme hydroxymethylbilane synthase (HMBS) as a crystal was fed substrate via a flow cell using Laue data from the ESRF. From these the putative binding site for the substrate has been experimentally determined and confirmed. The time course in the crystal was slower than expected. The most elongated electron density feature at the active site was seen at the 2 hour time point. By which time the initially colourless crystals of the active enzyme had become deep red characteristic of released cyclised HMB. In the second study a computer linear interpolation between the apocrustacyanin A1 subunit structure and A1 in the beta-crustacyanin (A1 with A3 with two bound astaxanthins) was made. This cartoon gives insights into the conformational changes of the A1 protein and carotenoid on binding. These changes are accompanied by a colour change of the carotenoid from orange red to blue/black. In a 3^rd study molecular dynamics of sugar binding to concanavalin A and to peanut lectin would be presented if time were available.