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Elucidating sequence, structural and dynamic basis of the functional regulation of membrane proteins

Award Number: R35GM142745
ORGANIZATION: NATIONAL INSTITUTE OF GENERAL MEDICAL SCIENCES
OPDIV: NIH
AWARD CLASS: DISCRETIONARY
AWARD ACTIVITY TYPE: SCIENTIFIC/HEALTH RESEARCH (INCLUDES SURVEYS)
PERIOD OF PERFORMANCE START DATE: 09/15/2021
PERIOD OF PERFORMANCE END DATE: 08/31/2026

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Elucidating sequence, structural and dynamic basis of the functional regulation of membrane proteins - Project Summary The overall aim of the research is Shukla group is to develop computational methods that facilitate investigation of rare conformational transitions in proteins and help guide the design of experiments to validate the in silico predictions. In par- ticular, we apply these computational methods to investigate functional regulation of membrane proteins such as membrane transporters and G-protein coupled receptors (GPCRs). Here, we propose development of transfer learning based methods to predict the effect of mutations on protein function and apply these methods to investigate monoamine transporters, sugar transporters and Class C GPCRs. Deep mutagenesis, whereby tens of thousands of mutational effects are determined by combining in vitro selections of sequence variants with Illumina sequencing, is an emerging technology for indirectly interrogating and observing protein conformations in living cells; the solving of an integrative structure of a neuronal class C G protein-coupled receptor in an active conformation by deep mutagenesis-guided modeling is one prominent example of this approach's success. Using deep mutagenesis and molecular dynamics simulations to inform each other, we plan to determine the mechanism of ion- coupled neurotransmitter import by monoamine transporters at atomic resolution. Fluorescent substrates have enabled us to use ﬂuorescence-based sorting of libraries of transporter mutants to ﬁnd mutations along the entire permeation pathway that increase or decrease substrate import. These comprehensive mutational landscapes will be used to interpret and support/reject hypotheses from simulations, including the role of ion-coupling in substrate transport regulation, proposed free energy barriers in the conformational-free energy landscape that limit import kinetics, and how sodium-neurotransmitter symport is coupled by a shared cytosolic exit pathway. Other notable features that arise from the deep mutational scans (e.g. putative regulatory sites) will be further explored, and a machine learning algorithm will be applied to transfer mutagenesis information to related transporters; the predicted mutational landscapes will then be validated by a small number of informative targeted mutants. We will further relate sequence to conformation and activity in metabotropic neurotransmitter receptors and sugar transporters. Finally, we plan to improve the proposed transfer algorithms by using deep learning techniques, which will facilitate integration of features derived from simulation datasets and multiple deep mutational scans to inform the effect of mutations on related proteins or tasks. The success of the proposed research program of results will be measured by development of algorithms that can accurately predict the variant effects on protein structure and function, elucidation of the mechanisms of ion-coupled regulation of neurotransmitter transport, selectivity mechanisms in sugar transporters and activation mechanisms of class C GPCRs.


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: 2026 ( Subtotal = $0 )
2026	2025	UNIVERSITY OF ILLINOIS	506 S WRIGHT ST	URBANA	IL	61801	CHAMPAIGN	USA	Biomedical Research and Research Training	000	5	11/21/2025	NON-COMPETING CONTINUATION	$0
														Subtotal = $0

Issue Date FY: 2025 ( Subtotal = $356,066 )
2025	2025	UNIVERSITY OF ILLINOIS	506 S WRIGHT ST	URBANA	IL	61801	CHAMPAIGN	USA	Biomedical Research and Research Training	000	5	9/9/2025	NON-COMPETING CONTINUATION	$356,066
														Subtotal = $356,066

Issue Date FY: 2024 ( Subtotal = $357,243 )
2024	2024	UNIVERSITY OF ILLINOIS	506 S WRIGHT ST	URBANA	IL	61801	CHAMPAIGN	USA	Biomedical Research and Research Training	000	4	9/11/2024	NON-COMPETING CONTINUATION	$357,243
														Subtotal = $357,243

Issue Date FY: 2023 ( Subtotal = $352,035 )
2023	2023	UNIVERSITY OF ILLINOIS	506 S WRIGHT ST	URBANA	IL	61801	CHAMPAIGN	USA	Biomedical Research and Research Training	000	3	9/20/2023	NON-COMPETING CONTINUATION	$352,035
														Subtotal = $352,035

Issue Date FY: 2022 ( Subtotal = $353,330 )
2022	2022	UNIVERSITY OF ILLINOIS	506 S WRIGHT ST	URBANA	IL	61801	CHAMPAIGN	USA	Biomedical Research and Research Training	000	2	9/26/2022	NON-COMPETING CONTINUATION	$353,330
														Subtotal = $353,330

Issue Date FY: 2021 ( Subtotal = $354,588 )
2021	2021	UNIVERSITY OF ILLINOIS	352 HENRY ADMINISTRATION BLDG	URBANA	IL	61801	CHAMPAIGN	USA	Biomedical Research and Research Training	000	1	9/14/2021	NEW	$354,588
														Subtotal = $354,588

Grand Total All Awards = $1,773,262

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Elucidating sequence, structural and dynamic basis of the functional regulation of membrane proteins

Award Number: R35GM142745

ORGANIZATION: NATIONAL INSTITUTE OF GENERAL MEDICAL SCIENCES

OPDIV: NIH

AWARD CLASS: DISCRETIONARY

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

PERIOD OF PERFORMANCE START DATE: 09/15/2021

PERIOD OF PERFORMANCE END DATE: 08/31/2026

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