title: Biomolecular dynamics studied with IR-spectroscopy using quantum cascade lasers combined with nanosecond perturbation techniques.
abstract: Early events of protein folding can be studied with fast perturbation techniques triggering non-equilibrium relaxation dynamics. A nanosecond laser-excited pH-jump or temperature-jump (T-jump) was applied to initiate helix folding or unfolding of poly-l-glutamic acid (PGA). PGA is a homopolypeptide with titratable carboxyl side-chains whose protonation degree determines the PGA conformation. A pH-jump was realized by the photochemical release of protons and induces PGA folding due to protonation of the side-chains. Otherwise, the helical conformation can be unfolded by a T-jump. We operated under conditions where PGA does not aggregate and temperature and pH are the regulatory properties of its conformation. The experiments were performed in such a manner that the folding/unfolding jump proceeded to the same PGA conformation. We quantified the increase/decrease in helicity induced by the pH-/T-jump and demonstrated that the T-jump results in a relatively small change in helical content in contrast to the pH-jump. This is caused by the strong pH-dependence of the PGA conformation. The conformational changes were detected by time-resolved single wavelength IR-spectroscopy using quantum cascade lasers (QCL). We could independently observe the kinetics for α-helix folding and unfolding in PGA by using different perturbation techniques and demonstrate the high sensitivity of time-resolved IR-spectroscopy to study protein folding mechanisms.
authors.affiliation: Department of Chemistry, University of Konstanz, 78457 Konstanz, Germany.
authors.forename: Alexander
authors.initials: A
authors.lastname: Popp
authors.affiliation: Department of Chemistry, University of Konstanz, 78457 Konstanz, Germany.
authors.forename: David
authors.initials: D
authors.lastname: Scheerer
authors.affiliation: Department of Chemistry, University of Konstanz, 78457 Konstanz, Germany.
authors.forename: Benjamin
authors.initials: B
authors.lastname: Heck
authors.affiliation: Department of Chemistry, University of Konstanz, 78457 Konstanz, Germany. Electronic address: karin.hauser@uni-konstanz.de.
authors.forename: Karin
authors.initials: K
authors.lastname: Hauser
mesh_terms: D006863:Hydrogen-Ion Concentration; D007700:Kinetics; D054023:Lasers, Semiconductor; D036103:Nanotechnology; D011099:Polyglutamic Acid / Q000737:chemistry; D011487:Protein Conformation; D017510:Protein Folding; D011506:Proteins / Q000032:analysis* / Q000737:chemistry; D013055:Spectrophotometry, Infrared / Q000295:instrumentation* / Q000379:methods*; D013696:Temperature
chemical_list: D011506:Proteins; D011099:Polyglutamic Acid
keywords: Folding kinetics; Infrared spectroscopy; Poly-l-glutamic acid; Quantum cascade laser; T-jump; pH-Jump