title: Folding of the SARS coronavirus spike glycoprotein immunological fragment (SARS_S1b): thermodynamic and kinetic investigation correlating with three-dimensional structural modeling.
abstract: Spike glycoprotein of SARS coronavirus (S protein) plays a pivotal role in SARS coronavirus (SARS_CoV) infection. The immunological fragment of the S protein (Ala251-His641, SARS_S1b) is believed to be essential for SARS_CoV entering the host cell through S protein-ACE-2 interaction. We have quantitatively characterized the thermally induced and GuHCl-induced unfolding features of SARS_S1b using circular dichroism (CD), tryptophan fluorescence, and stopped-flow spectral techniques. For the thermally induced unfolding at pH 7.4, the apparent activation energy (E(app)) and transition midpoint temperature (Tm) were determined to be 16.3 +/- 0.2 kcal/mol and 52.5 +/- 0.4 degrees C, respectively. The CD spectra are not dependent on temperature, suggesting that the secondary structure of SARS_S1b has a relatively high thermal stability. GuHCl strongly affected SARS_S1b structure. Both the CD and fluorescent spectra resulted in consistent values of the transition middle concentration of the denaturant (Cm, ranging from 2.30 to 2.45 M) and the standard free energy change (deltaG(o), ranging from 2.1 to 2.5 kcal/mol) for the SARS_S1b unfolding reaction. Moreover, the kinetic features of the chemical unfolding and refolding of SARS_S1b were also characterized using a stopped-flow CD spectral technique. The obvious unfolding reaction rates and relaxation times were determined at various GuHCl concentrations, and the Cm value was obtained, which is very close to the data that resulted from CD and fluorescent spectral determinations. Secondary and three-dimensional structural predictions by homology modeling indicated that SARS_S1b folded as a globular-like structure by beta-sheets and loops; two of the four tryptophans are located on the protein surface, which is in agreement with the tryptophan fluorescence result. The three-dimensional model was also used to explain the recently published experimental results of S1-ACE-2 binding and immunizations.
authors.affiliation: Drug Discovery and Design Center, State Key Laboratory of Drug Research, Shanghai Institute of Materia Medica, Shanghai Institutes for Biological Sciences, Graduate School, Chinese Academy of Sciences, Shanghai 201203, China.
authors.forename: Changying
authors.initials: C
authors.lastname: Yu
authors.forename: Chunshan
authors.initials: C
authors.lastname: Gui
authors.forename: Haibin
authors.initials: H
authors.lastname: Luo
authors.forename: Lili
authors.initials: L
authors.lastname: Chen
authors.forename: Liang
authors.initials: L
authors.lastname: Zhang
authors.forename: Hao
authors.initials: H
authors.lastname: Yu
authors.forename: Sheng
authors.initials: S
authors.lastname: Yang
authors.forename: Weihong
authors.initials: W
authors.lastname: Jiang
authors.forename: Jianhua
authors.initials: J
authors.lastname: Shen
authors.forename: Xu
authors.initials: X
authors.lastname: Shen
authors.forename: Hualiang
authors.initials: H
authors.lastname: Jiang
mesh_terms: D000595:Amino Acid Sequence; D002942:Circular Dichroism; D003198:Computer Simulation; D019791:Guanidine / Q000737:chemistry; D006358:Hot Temperature; D007700:Kinetics; D008562:Membrane Glycoproteins / Q000737:chemistry* / Q000276:immunology; D008956:Models, Chemical; D008958:Models, Molecular; D008969:Molecular Sequence Data; D010446:Peptide Fragments / Q000737:chemistry* / Q000276:immunology; D011489:Protein Denaturation; D017510:Protein Folding; D017433:Protein Structure, Secondary; D045473:Severe acute respiratory syndrome-related coronavirus / Q000737:chemistry*; D013050:Spectrometry, Fluorescence; D064370:Spike Glycoprotein, Coronavirus; D040681:Structural Homology, Protein; D013816:Thermodynamics; D014364:Tryptophan / Q000737:chemistry; D014759:Viral Envelope Proteins / Q000737:chemistry* / Q000276:immunology
chemical_list: D008562:Membrane Glycoproteins; D010446:Peptide Fragments; D064370:Spike Glycoprotein, Coronavirus; D014759:Viral Envelope Proteins; C578557:spike glycoprotein, SARS-CoV; C578553:spike protein, mouse hepatitis virus; D014364:Tryptophan; D019791:Guanidine