title: Structural characterization of Hsp12, the heat shock protein from Saccharomyces cerevisiae, in aqueous solution where it is intrinsically disordered and in detergent micelles where it is locally α-helical.
abstract: Hsp12 (heat shock protein 12) belongs to the small heat shock protein family, partially characterized as a stress response, stationary phase entry, late embryonic abundant-like protein located at the plasma membrane to protect membrane from desiccation. Here, we report the structural characterization of Hsp12 by NMR and biophysical techniques. The protein was labeled uniformly with nitrogen-15 and carbon-13 so that its conformation could be determined in detail both in aqueous solution and in two membrane-mimetic environments, SDS and dodecylphosphocholine (DPC) micelles. Secondary structural elements determined from assigned chemical shifts indicated that Hsp12 is dynamically disordered in aqueous solution, whereas it gains four helical stretches in the presence of SDS micelles and a single helix in presence of DPC. These conclusions were reinforced by circular dichroism spectra of the protein in all three environments. The lack of long range interactions in NOESY spectra indicated that the helices present in SDS micelles do not pack together. R(1) and R(2), relaxation and heteronuclear NOE measurements showed that the protein is disordered in aqueous solution but becomes more ordered in presence of detergent micelles. NMR spectra collected in presence of paramagnetic spin relaxation agents (5DSA, 16DSA, and Gd(DTPA-BMA)) indicated that the amphipathic α-helices of Hsp12 in SDS micelles lie on the membrane surface. These observations are in agreement with studies suggesting that Hsp12 functions to protect the membrane from desiccation.
authors: National Magnetic Resonance Facility at Madison, University of Wisconsin, Madison, Wisconsin 53705, USA.
authors: Kiran K
authors: KK
authors: Singarapu
authors: Marco
authors: M
authors: Tonelli
authors: Darius C
authors: DC
authors: Chow
authors: Ronnie O
authors: RO
authors: Frederick
authors: William M
authors: WM
authors: Westler
authors: John L
authors: JL
authors: Markley
mesh_terms: D006360:Heat-Shock Proteins / Q000737:chemistry*; D009682:Magnetic Resonance Spectroscopy; D008823:Micelles; D010767:Phosphorylcholine / Q000031:analogs & derivatives / Q000737:chemistry; D017433:Protein Structure, Secondary; D029701:Saccharomyces cerevisiae Proteins / Q000737:chemistry*; D012967:Sodium Dodecyl Sulfate / Q000737:chemistry
chemical_list: C112591:HSP12 protein, S cerevisiae; D006360:Heat-Shock Proteins; D008823:Micelles; D029701:Saccharomyces cerevisiae Proteins; D010767:Phosphorylcholine; D012967:Sodium Dodecyl Sulfate; C028810:dodecylphosphocholine