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Title: Local Crowd, Local Probe: Strengths and Drawbacks of Azidohomoalanine as a Site-Specific Crowding Probe
Every residue on a protein can be characterized by its interaction with water, in lack or in excess, as water is the matrix of biological systems. Infrared spectroscopy and the implementation of local azidohomoalanine (AHA) probes allow us to move beyond an ensemble or surface-driven conceptualization of water behavior and toward a granular, site-specific picture. In this paper, we examined the role of crowding in modulating both global and local behavior on the β-hairpin, TrpZip2 using a combination of Fourier-transform infrared spectroscopy (FTIR) spectroscopy, two-dimensional infrared (2D IR) spectroscopy, and molecular dynamics simulations. We found that, at the amino acid level, crowding drove dehydration of both sheet and turn peptide sites as well as free AHA. However, the subpicosecond dynamics showed highly individualized responses based on the local environment. Interestingly, while steady-state FTIR measurements revealed similar responses at the amino-acid level to hard versus soft crowding (dehydration), we found that PEG and glucose had opposite stabilizing and destabilizing effects on the protein secondary structure, emphasizing an important distinction in understanding the impact of crowding on protein structure as well as the role of crowding across length scales.  more » « less
Award ID(s):
1847199
PAR ID:
10532365
Author(s) / Creator(s):
;
Publisher / Repository:
American Chemical Society
Date Published:
Journal Name:
The Journal of Physical Chemistry B
Volume:
128
Issue:
22
ISSN:
1520-6106
Page Range / eLocation ID:
5310 to 5319
Format(s):
Medium: X
Sponsoring Org:
National Science Foundation
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