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Conformational Studies of the Region of Influenza A M2 Protein Involved in Viral Budding

Award Number: R15AI117657
ORGANIZATION: NATIONAL INSTITUTE OF ALLERGY & INFECTIOUS DISEASES
OPDIV: NIH
AWARD CLASS: DISCRETIONARY
AWARD ACTIVITY TYPE: SCIENTIFIC/HEALTH RESEARCH (INCLUDES SURVEYS)
PERIOD OF PERFORMANCE START DATE: 09/01/2015
PERIOD OF PERFORMANCE END DATE: 06/30/2019

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Conformational Studies of the Region of Influenza A M2 Protein Involved in Viral Budding - ¿ DESCRIPTION (provided by applicant): Influenza A virus is a major public health concern because of its annual death toll and its potential to cause devastating pandemics. Fundamental information on the life cycle of the influenza virus is still being discovered which could lead to new tactics for treating influenza infections. Recently a series of studies on the role of the influenza A M2 protein in virus assembly and budding have been published. This proposal describes experiments designed to understand the conformation and dynamics of the C-terminal juxtamembrane region of the M2 protein shown to be critical to viral budding. The majority of previously published biophysical work on M2 has focused on truncated M2 peptide constructs solubilized in detergent or in bilayers of limited physiological relevance. The full-length protein can now be efficiently over-expressed and purified. To eliminate potential artifacts due to the use of fragments of M2 and distortion of the structure due to non-native hydrophobic environments, the work proposed here will use full-length protein reconstituted into membrane bilayers that reflect the lipid composition of influenza A virions as well as model membrane bilayers in which viral budding functional assays have been carried out. An atomic level understanding of the role M2 plays in viral budding requires detailed structural information. Our strategy for characterizing the structure and dynamics of the C-terminal juxtamembrane region of the full-length influenza A M2 protein will rely on site-directed spin label (SDSL) EPR experiments. We will prepare a series of full-length M2 proteins with spin labels in the C- region (residues 46-62) shown previously to play an essential role in viral budding. SDSL EPR data used in structure determination will include patterns of spin label mobilities, distance constraint from spin-spin couplings and accessibility to paramagnetic reagents of varying bilayer/aqueous solubility. Using deuterated forms of cholesterol, electron spin-echo envelope modulation (ESEEM) experiments will be used to measure distances between the deuterium nuclei on cholesterol and spin-labeled sites on the M2 protein. The high sensitivity of SDSL EPR studies will allow a range of cholesterol:peptide:lipid ratios to be tested, enabling a fuller characterization of the location(s) and affinities of cholesterol binding than has been previously possible.


Issue Date FY	Funding FY	Legal Entity Name	Legal Entity Address	Legal Entity City	Legal Entity State	Legal Entity Zip Code	Legal Entity COUNTY	Legal Entity COUNTRY	Assistance Listing	Award Code	Budget Year	Action Date	Action Type	Action Amount

Issue Date FY: 2021 ( Subtotal = $0 )
2021	2015	SWARTHMORE COLLEGE	500 COLLEGE AVE	SWARTHMORE	PA	19081	DELAWARE	USA	Allergy and Infectious Diseases Research	000	1	10/1/2020	NEW	$0
2021	2015	SWARTHMORE COLLEGE	500 COLLEGE AVE	SWARTHMORE	PA	19081	DELAWARE	USA	Allergy and Infectious Diseases Research	001	1	10/3/2020	NEW	$0
2021	2015	SWARTHMORE COLLEGE	500 COLLEGE AVE	SWARTHMORE	PA	19081	DELAWARE	USA	Allergy and Infectious Diseases Research	002	1	10/3/2020	NEW	$0
														Subtotal = $0

Issue Date FY: 2019 ( Subtotal = -$180 )
2019	2015	SWARTHMORE COLLEGE	500 COLLEGE AVE	SWARTHMORE	PA	19081	DELAWARE	USA	Allergy and Infectious Diseases Research	000	1	8/28/2019	NEW	-$180
														Subtotal = -$180

Issue Date FY: 2015 ( Subtotal = $326,754 )
2015	2015	SWARTHMORE COLLEGE	500 COLLEGE AVENUE	SWARTHMORE	PA	19081	DELAWARE	USA	Allergy and Infectious Diseases Research	000	1	8/25/2015	NEW	$326,754
														Subtotal = $326,754

Grand Total All Awards = $326,574

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Conformational Studies of the Region of Influenza A M2 Protein Involved in Viral Budding

Award Number: R15AI117657

ORGANIZATION: NATIONAL INSTITUTE OF ALLERGY & INFECTIOUS DISEASES

OPDIV: NIH

AWARD CLASS: DISCRETIONARY

AWARD ACTIVITY TYPE: SCIENTIFIC/HEALTH RESEARCH (INCLUDES SURVEYS)

PERIOD OF PERFORMANCE START DATE: 09/01/2015

PERIOD OF PERFORMANCE END DATE: 06/30/2019

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