Official websites use .gov

A .gov website belongs to an official government organization in the United States.

Secure .gov websites use HTTPS

A lock ( 🔒 ) or https:// means you've safely connected to the .gov website. Share sensitive information only on official, secure websites.

Need Help

Integrative structural analysis of the vRNAP-coupled Coliphage N4 ejectosome

Award Number: F31AI194663
ORGANIZATION: NATIONAL INSTITUTE OF ALLERGY & INFECTIOUS DISEASES
OPDIV: NIH
AWARD CLASS: DISCRETIONARY
AWARD ACTIVITY TYPE: FELLOWSHIP/SCHOLARSHIP/STUDENT LOANS
PERIOD OF PERFORMANCE START DATE: 02/01/2027
PERIOD OF PERFORMANCE END DATE: 01/31/2028

Group Awards By:

View Award Description

Integrative structural analysis of the vRNAP-coupled Coliphage N4 ejectosome - ABSTRACT Bacteriophage N4, the model virus of the newly established and understudied Schitoviridae family, employs a unique mechanism to translocate its genome into Gram-negative bacterial hosts. This process is mediated by gp50, a massive 380 kDa virion-associated RNA polymerase (vRNAP). N4 vRNAP's role in initiating transcription immediately after infection is well studied; however, its role in forming a specialized membrane- spanning complex termed the ejectosome is less understood. The transcriptional activity of this protein is carried out by the middle 1100 residues; however, domains of equal or greater length surround the transcriptional core on both the N and C termini, and these domains remain largely uncharacterized structurally and functionally. I have recently determined structures of the gp50 RNAP and C-terminal domains (currently under review), but the N-terminal domain and the complete ejectosome complex remain unresolved. This proposal addresses these knowledge gaps by leveraging state-of-the-art cryogenic electron microscopy (Cryo- EM) and electron tomography (Cryo-ET) approaches. I hypothesize that multiple copies of vRNAP (gp50) are structurally integrated into a membrane-spanning channel formed by N4 ejection proteins gp51 and gp52, with the N-terminal domain (NTD) of vRNAP contributing to the channel structure while the RNAP and C-terminal domains remain tethered to facilitate DNA translocation. In Aim 1, I will reconstitute the complete ejectosome complex in vitro from purified components (gp50-NTD, gp51, and gp52) and determine its high-resolution structure using Cryo-EM single particle analysis, revealing the structure of the N-terminal domain and its integration into the membrane-spanning channel. In Aim 2, I will use in situ Cryo-ET to visualize the N4 ejectosome in its pre- and post-ejection states. By analyzing mature virions and N4-infected bacterial minicells, I will capture the internal organization of vRNAP and associated ejection proteins before and after translocation. These experiments will provide insight into the structural transitions required for ejectosome assembly and function during early infection. Together, these studies will advance our understanding of the molecular mechanisms underlying genome ejection in N4 and related phages. Understanding the structural basis of genome ejection in this therapeutically relevant phage family has implications for phage therapy development, rational engineering of phage-based delivery systems, and microbiome dynamics. This work will provide me with comprehensive training in integrative structural biology approaches, including advanced Cryo- EM and Cryo-ET techniques, preparing me for an independent research career.


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 = $43,769 )
2026	2026	UNIVERSITY OF ALABAMA AT BIRMINGHAM	701 S 20TH STREET	BIRMINGHAM	AL	35294	JEFFERSON	USA	Allergy and Infectious Diseases Research	000	1	8/1/2026	NEW	$43,769
														Subtotal = $43,769

Grand Total All Awards = $43,769

Top

All Categories

About

Search

Reports

Data Submission

Award Information

Integrative structural analysis of the vRNAP-coupled Coliphage N4 ejectosome

Award Number: F31AI194663

ORGANIZATION: NATIONAL INSTITUTE OF ALLERGY & INFECTIOUS DISEASES

OPDIV: NIH

AWARD CLASS: DISCRETIONARY

AWARD ACTIVITY TYPE: FELLOWSHIP/SCHOLARSHIP/STUDENT LOANS

PERIOD OF PERFORMANCE START DATE: 02/01/2027

PERIOD OF PERFORMANCE END DATE: 01/31/2028

Federal Websites

Department of Health & Human Services

HHS Operating Divisions

HHS Staff Divisions

Download A Document Viewer