The study of the bacterial mechanosensitive MscL channel has biomedical significance for several reasons. First, the channel serves a vital function in maintaining osmotic homeostasis of microbes;when the channel misfunctions it can lead to the death of the microbial cell, and thus may be a viable pharmacological target. Second, as nanotechnology progresses, the potential for biological sensors, especially MscL, to be used in biomedical nano-devices is being realized. Third, MscL is an excellent model for the study of mechanosensory transduction. Because of its tractable nature and the existence of a crystal structure, MscL is a unique tool to investigate structure-function relationships in a mechanosensory channel. We have been exploiting this system to obtain a better understanding of the molecular mechanisms of how a mechanosensitive channel senses and responds to membrane tension;this is the long-term objective of this proposal. While models for structural transitions during gating have been proposed, they are not consistent or complete, and many of the fundamental features are not yet resolved. The experiments within this proposal are designed to ally the solved structure with molecular, biochemical and electrophysiological analyses to determine the functional role that regions of the protein play in sensing and responding to membrane stretch, to define transitions that occur upon gating, and to determine aspects of the open-pore structure. The approaches used include: the generation of chimeras to determine the structural elements associated with functional differences of homologues, utilizing the results from a sulfhydryl-based post-translational scan to determine residues that should be further tested by mutagenesis for whether they enter or exit a lipid environment upon gating, disulfide trapping to define closed, transition and open states of the channel, and attempts to crystallize the channel in these multiple states.

Public Health Relevance

Studying how a bacterial sensor detects forces will allow insight into the mechanisms of how human mechanosensors, e.g. those used in blood pressure and kidney regulation, may function;thus, we may eventually speculate how such sensors can be modulated by drugs. This work could also have implications in anti-bacterial drug design and the utilization of biological sensors for future technological feats, such as nano- devices for drug delivery.

Agency
National Institute of Health (NIH)
Institute
National Institute of General Medical Sciences (NIGMS)
Type
Research Project (R01)
Project #
5R01GM061028-12
Application #
8538996
Study Section
Biophysics of Neural Systems Study Section (BPNS)
Program Officer
Nie, Zhongzhen
Project Start
2000-06-01
Project End
2015-08-31
Budget Start
2013-09-01
Budget End
2014-08-31
Support Year
12
Fiscal Year
2013
Total Cost
$322,214
Indirect Cost
$119,564
Name
University of Texas Sw Medical Center Dallas
Department
Physiology
Type
Schools of Medicine
DUNS #
800771545
City
Dallas
State
TX
Country
United States
Zip Code
75390
Wray, Robin; Iscla, Irene; Gao, Ya et al. (2016) Dihydrostreptomycin Directly Binds to, Modulates, and Passes through the MscL Channel Pore. PLoS Biol 14:e1002473
Malcolm, Hannah R; Blount, Paul (2015) Mutations in a Conserved Domain of E. coli MscS to the Most Conserved Superfamily Residue Leads to Kinetic Changes. PLoS One 10:e0136756
Malcolm, Hannah R; Blount, Paul; Maurer, Joshua A (2015) The mechanosensitive channel of small conductance (MscS) functions as a Jack-in-the box. Biochim Biophys Acta 1848:159-66
Iscla, Irene; Wray, Robin; Eaton, Christina et al. (2015) Scanning MscL Channels with Targeted Post-Translational Modifications for Functional Alterations. PLoS One 10:e0137994
Iscla, Irene; Wray, Robin; Blount, Paul et al. (2015) A new antibiotic with potent activity targets MscL. J Antibiot (Tokyo) 68:453-62
Zhong, Dalian; Blount, Paul (2014) Electrostatics at the membrane define MscL channel mechanosensitivity and kinetics. FASEB J 28:5234-41
Iscla, Irene; Wray, Robin; Wei, Shuguang et al. (2014) Streptomycin potency is dependent on MscL channel expression. Nat Commun 5:4891
Zhong, Dalian; Yang, Li-Min; Blount, Paul (2014) Dynamics of protein-protein interactions at the MscL periplasmic-lipid interface. Biophys J 106:375-81
Zhong, Dalian; Blount, Paul (2013) Phosphatidylinositol is crucial for the mechanosensitivity of Mycobacterium tuberculosis MscL. Biochemistry 52:5415-20
Iscla, Irene; Eaton, Christina; Parker, Juandell et al. (2013) Improving the Design of a MscL-Based Triggered Nanovalve. Biosensors (Basel) 3:171-84

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