title: Pre-denaturing transitions in human serum albumin probed using time-resolved phosphorescence.
abstract: The investigation of protein dynamics has long been of interest, since protein interactions and functions can be determined by their structure and changes in conformation. Although fluorescence, occurring on the nanosecond timescale, from intrinsic fluorescent amino acids has been extensively used, in order to fully access conformational changes longer timescales are required. Phosphorescence enables processes on the microsecond to second timescale to be accessed. However, at room temperature this emission can be weak and non trivial to measure. It requires the removal of oxygen - a common triplet state quencher and appropriate instrumentation. In this work we make use of a chemical deoxygenator to study room temperature phosphorescence from tryptophan in human serum albumin excited using a pulsed UV light emitting diode. This is extended to monitor the phosphorescence emission upon increasing temperature, allowing pre-denaturing transitions to be observed. Time-resolved data are analysed, both as the sum of exponential decays and using a distribution analysis based on non extensive decay kinetics. These results are compared to a fluorescence study and both the average lifetime and contribution of the different emitting components were found to give more dramatic changes on the phosphorescence timescale.
authors: HORIBA Jobin Yvon IBH Ltd., 45 Finnieston Street, Glasgow G3 8JU, UK.
authors: Kulwinder
authors: K
authors: Sagoo
authors: HORIBA Jobin Yvon IBH Ltd., 45 Finnieston Street, Glasgow G3 8JU, UK.
authors: Richard
authors: R
authors: Hirsch
authors: HORIBA Jobin Yvon IBH Ltd., 45 Finnieston Street, Glasgow G3 8JU, UK.
authors: Pamela
authors: P
authors: Johnston
authors: HORIBA Jobin Yvon IBH Ltd., 45 Finnieston Street, Glasgow G3 8JU, UK.
authors: David
authors: D
authors: McLoskey
authors: HORIBA Jobin Yvon IBH Ltd., 45 Finnieston Street, Glasgow G3 8JU, UK. Electronic address: graham.hungerford@horiba.com.
authors: Graham
authors: G
authors: Hungerford
mesh_terms: D006801:Humans; D049449:Luminescence; D011489:Protein Denaturation; D012709:Serum Albumin / Q000378:metabolism*; D013050:Spectrometry, Fluorescence; D013696:Temperature; D013997:Time Factors
chemical_list: D012709:Serum Albumin
keywords: Distribution analysis; Fluorescence lifetime; Non extensive decay; Protein; Room temperature phosphorescence