Protein-ice interaction of an antifreeze protein observed with solid-state NMR.
NMR on frozen solutions is an ideal method to study fundamental questions of macromolecular hydration, because the hydration shell of many biomolecules does not freeze together with bulk solvent. In the present study, we present previously undescribed NMR methods to study the interactions of proteins with their hydration shell and the ice lattice in frozen solution. We applied these methods to compare solvent interaction of an ice-binding type III antifreeze protein (AFP III) and ubiquitin a non-ice-binding protein in frozen solution. We measured (1)H-(1)H cross-saturation and cross-relaxation to provide evidence for a molecular contact surface between ice and AFP III at moderate freezing temperatures of -35 °C. This phenomenon is potentially unique for AFPs because ubiquitin shows no such cross relaxation or cross saturation with ice. On the other hand, we detected liquid hydration water and strong water-AFP III and water-ubiquitin cross peaks in frozen solution using relaxation filtered (2)H and HETCOR spectra with additional (1)H-(1)H mixing. These results are consistent with the idea that ubiquitin is surrounded by a hydration shell, which separates it from the bulk ice. For AFP III, the water cross peaks indicate that only a portion of its hydration shell (i.e., at the ice-binding surface) is in contact with the ice lattice. The rest of AFP III's hydration shell behaves similarly to the hydration shell of non-ice-interacting proteins such as ubiquitin and does not freeze together with the bulk water.
Siemer
Ansgar B
AB
Department of Chemistry, Columbia University, 3000 Broadway, New York, NY 10027, USA.
Huang
Kuo-Ying
KY
McDermott
Ann E
AE
D021343:Antifreeze Proteins, Type III / Q000378:metabolism*; D002247:Carbon Isotopes / Q000378:metabolism; D007053:Ice; D008956:Models, Chemical; D009587:Nitrogen Isotopes / Q000378:metabolism; D019906:Nuclear Magnetic Resonance, Biomolecular / Q000379:methods*; D011522:Protons; D025801:Ubiquitin / Q000378:metabolism; D014867:Water / Q000378:metabolism
D021343:Antifreeze Proteins, Type III; D002247:Carbon Isotopes; D007053:Ice; D009587:Nitrogen Isotopes; D011522:Protons; D025801:Ubiquitin; D014867:Water