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Int. J. Mol. Sci. 2013, 14(9), 18881-18898; doi:10.3390/ijms140918881

Spectroscopic Studies of Model Photo-Receptors: Validation of a Nanosecond Time-Resolved Micro-Spectrophotometer Design Using Photoactive Yellow Protein and α-Phycoerythrocyanin

Department of Physics, University of Wisconsin-Milwaukee, 1900 E. Kenwood Blvd., Milwaukee, WI 53211, USA
These authors contributed equally to this work.
Author to whom correspondence should be addressed.
Received: 22 July 2013 / Revised: 27 August 2013 / Accepted: 2 September 2013 / Published: 13 September 2013
(This article belongs to the Special Issue Frontiers of Micro-Spectroscopy in Biological Applications)
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Time-resolved spectroscopic experiments have been performed with protein in solution and in crystalline form using a newly designed microspectrophotometer. The time-resolution of these experiments can be as good as two nanoseconds (ns), which is the minimal response time of the image intensifier used. With the current setup, the effective time-resolution is about seven ns, determined mainly by the pulse duration of the nanosecond laser. The amount of protein required is small, on the order of 100 nanograms. Bleaching, which is an undesirable effect common to photoreceptor proteins, is minimized by using a millisecond shutter to avoid extensive exposure to the probing light. We investigate two model photoreceptors, photoactive yellow protein (PYP), and α-phycoerythrocyanin (α-PEC), on different time scales and at different temperatures. Relaxation times obtained from kinetic time-series of difference absorption spectra collected from PYP are consistent with previous results. The comparison with these results validates the capability of this spectrophotometer to deliver high quality time-resolved absorption spectra.
Keywords: nanosecond spectroscopy; time-resolved spectroscopy; photoreceptor proteins nanosecond spectroscopy; time-resolved spectroscopy; photoreceptor proteins

This is an open access article distributed under the Creative Commons Attribution License (CC BY 3.0).

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Purwar, N.; Tenboer, J.; Tripathi, S.; Schmidt, M. Spectroscopic Studies of Model Photo-Receptors: Validation of a Nanosecond Time-Resolved Micro-Spectrophotometer Design Using Photoactive Yellow Protein and α-Phycoerythrocyanin. Int. J. Mol. Sci. 2013, 14, 18881-18898.

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