DETERMINATION OF SOLUTION STRUCTURES OF PARAMAGNETIC PROTEINS BY NMR

Standard procedures for using nuclear Overhauser enhancements (NOE) be tween protons to generate structures for diamagnetic proteins in solut ion from NMR data may be supplemented by using dipolar shifts if the p rotein is paramagnetic. This is advantageous since the electron-nuclea r dipolar coupling provides relatively long-range geometric informatio n with respect to the paramagnetic centre which complements the short- range distance constraints from NOEs. Several different strategies hav e been developed to date, but none of these attempts to combine data f rom NOEs and dipolar shifts in the initial stages of structure calcula tion or to determine three dimensional protein structures together wit h their magnetic properties. This work shows that the magnetic and ato mic structures are highly correlated and that it is important to have additional constraints both to provide starting parameters for the mag netic properties and to improve the definition of the best fit. Useful parameters can be obtained for haem proteins from Fermi contact shift s; this approach is compared with a new method based on the analysis o f dipolar shifts in haem methyl groups with respect to data from horse and tuna ferricytochromes c. The methods developed for using data fro m NOEs and dipolar shifts have been incorporated in a new computer pro gram, PARADYANA, which is demonstrated in application to a model data set for the sequence of the haem octapeptide known as microperoxidase- 8.