Comparison of the solution and crystal structures of staphylococcal nuclease with 13C and 15N chemical shifts used as structural fingerprints.
We report high-resolution 13C and 15N NMR spectra of crystalline staphylococcal nuclease (Nase) complexed to thymidine 3',5'-diphosphate and Ca2+. High sensitivity and resolution are obtained by applying solid-state NMR techniques--high power proton decoupling and cross-polarization magic angle sample spinning (CPMASS)--to protein samples that have been efficiently synthesized and labeled by an overproducing strain of Escherichia coli. A comparison of CPMASS and solution spectra of Nase labeled with either [methyl-13C]methionine or [15N]valine shows that the chemical shifts in the crystalline and solution states are virtually identical. This result is strong evidence that the protein conformations in the solution and crystalline states are nearly the same. Because of the close correspondence of the crystal and solution chemical shifts, sequential assignments obtained in solution apply to the crystal spectra. It should therefore be possible to study the molecular structure and dynamics of many sequentially assigned atomic sites in Nase crystals. Similar experiments are applicable to the growing number of proteins that can be obtained from efficient expression systems.
Cole
H B
HB
Bone Research Branch, National Institute of Dental Research, Bethesda, MD 20892.
Sparks
S W
SW
Torchia
D A
DA
D002247:Carbon Isotopes; D003460:Crystallization; D009682:Magnetic Resonance Spectroscopy / Q000379:methods; D008715:Methionine; D008836:Micrococcal Nuclease / Q000378:metabolism*; D008958:Models, Molecular; D009587:Nitrogen Isotopes; D011487:Protein Conformation; D012996:Solutions; D014633:Valine
D002247:Carbon Isotopes; D009587:Nitrogen Isotopes; D012996:Solutions; D008715:Methionine; D008836:Micrococcal Nuclease; D014633:Valine