Difference: AneelAggarwal (10 vs. 11)
Revision 1101 Apr 2008 - Main.DavidCowburn
Public information on Grants associated with NYSBC Grant Number: 2R01AI041706-06 PI Name: AGGARWAL, ANEEL K. PI Email: aggarwal@inka.mssm.edu PI Title: PROFESSOR Project Title: THE ROLE OF IRF PROTEINS IN GENE TRANSCRIPTION Abstract: DESCRIPTION (provided by applicant): The IRF family of transcription factors plays critical roles in the regulation of interferons in response to viral infection, and in the development and functioning of the immune system. The IRF family includes now over ten members, characterized by a homologous DNA binding domain (DBD) at the N-terminus. We propose structural studies to explore the broader role of IRF-3 in the regulation of interferon-beta (IFN-beta) gene expression, and the role of IRF-4 in the development and functioning of the immune system. IRF-3 is activated in virally infected cells by phosphorylation of specific residues at the C-terminus, leading to nuclear translocation and binding to a so-called PRD I-III DNA element in the IFN-beta promoter. IRF-4 binds to a number of composite DNA elements in the promoters and enhancers of B-lymphoid and myeloid genes, but exclusively in association with PU.1. Specific aims are: 1) Determine the structure of IRF-3 DBD bound to the entire PRD I-III element by crystallographic methods. 2) Determine the structure of intact, phosphorylated IRF-3 bound to the PRD I-III element. IRF-3 will be phosphorylated in vitro prior to crystallization. 3) Structurally characterize an autoinhibitory element at the N-terminus of IRF-4 by NMR methods. 4) Determine the structure of IRF-4 in complex with phosphorylated PU.I. PU.1 will be phosphorylated in vitro by casein kinase II prior to crystallization. Together, these aims explore the specificity and cooperativity of these IRFs, and the role of phosphorylation and autoinhibitory elements. Thesaurus Terms: gene expression, gene induction /repression, genetic regulation, genetic transcription, interferon beta, protein isoform, protein structure function, transcription factor DNA binding protein, genetic regulatory element, immune system, immunity, phosphorylation, protein protein interaction crystallization, nuclear magnetic resonance spectroscopy Institution: MOUNT SINAI SCHOOL OF MEDICINE OF NYU OF NEW YORK UNIVERSITY NEW YORK, NY 10029 Fiscal Year: 2005 Department: PHYSIOLOGY AND BIOPHYSICS Project Start: 01-JUN-1998 Project End: 30-NOV-2009 ICD: NATIONAL INSTITUTE OF ALLERGY AND INFECTIOUS DISEASES IRG: ZRG1 Grant Number: 2R01GM062947-05 PI Name: AGGARWAL, ANEEL K. PI Email: aggarwal@inka.mssm.edu PI Title: PROFESSOR Project Title: STRUCTURE & ASSEMBLY OF TRANSLATION REPRESSOR COMPLEXES Abstract: DESCRIPTION (provided by applicant): Translation regulation plays a vital role in the lives of most organisms. It provides an important checkpoint in the pathways for cell growth and differentiation, and a link to the pathology of several diseases. A prominent example of translational regulation in early fly embryogenesis is the repression of maternal hunchback mRNA by Pumilio, Nanos, and Brain tumor. The Nanos protein is only synthesized at the posterior of the embryo due to the translational repression of its own maternal mRNA by Smaug. Pumilio and Nanos are also required for the translation repression of maternal Cyclin B mRNA. Our long-term objective is to uncover the structure and mechanism of assembly of these translational regulators. Specific aims are: 1. Test our basic hypothesis that two Pumilio molecules bind to a single hunchback regulatory element. This will accomplished through a combination of structure and genetics. 2. Identify specific residues in Brain tumor that interact with Nanos. 3. Cocrystallize Pumilio and Nanos in a ternary complex with hunchback mRNA. 4. Determine the structure of Pumilio Puf domain bound to Cyc B mRNA. We will first define the minimal element in the Cyclin B mRNA that binds Pumilio with high affinity and confers regulation in vivo. 5. Elucidate the structure of a SAM domain/RNA complex by NMR methods. 6. Guided by structure, isolate a Smaug dominant negative for the identification of potential corepressors. Together, these structural and genetic experiments will provide a molecular basis for the translation regulatory events that organize the body pattern along the anterior-posterior axis. Thesaurus Terms: There are no thesaurus terms on file for this project Institution: MOUNT SINAI SCHOOL OF MEDICINE OF NYU OF NEW YORK UNIVERSITY NEW YORK, NY 10029 Fiscal Year: 2005 Department: PHYSIOLOGY AND BIOPHYSICS Project Start: 01-MAY-2001 Project End: 30-APR-2009 ICD: NATIONAL INSTITUTE OF GENERAL MEDICAL SCIENCES IRG: ZRG1 Grant Number: 5R01GM044006-15 PI Name: AGGARWAL, ANEEL K. PI Email: aggarwal@inka.mssm.edu PI Title: PROFESSOR Project Title: RECOGNITION AND CLEAVAGE OF DNA BY RESTRICTION ENZYMES | ||||||||
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| < < | Abstract: DESCRIPTION (provided by applicant): Protein-DNA selectivity is a central event in many biological processes, ranging from transcription and replication to restriction and modification. Type II restriction endonucleases are ideal systems for studying selectivity because of their high specificity and great variety. The enzymes recognize and cleave DNA sequences that vary between four to eight base pairs. Their sequence specificity is remarkable. A single base pair change within the recognition sequence can lead to well over a million fold reduction in activity. An understanding of sequence specific cleavage is relevant to protein mediating site-specific recombination and DNA repair by excision. The long-term goals of this projectyiyiu are to understand the mechanism by which type II restriction enzymes recognize and cleave DNA, and to design mutants with altered specificities. The three broad aims are: 1) Determine the basis of discrimination between closely related DNA sites. With structures of BamHI? and Bglll in hand, we will now determine structures of an "intermediate" endonuclease with the specificity of both BamHI? and Bglll: BstYI. We will also manipulate the specificity of BamHI? with the aid of these new structures. 2) Determine the basis of specific versus non-specific DNA binding. We have determined the structure of BamHI? bound to one non-cognate DNA sequence. We will now determine structures of BamHI? bound to other non-cognate DNA sequences, in order to see the structural adaptations in going from one non-cognate sequence to another. We will also experimentally test a model of the non-cognate complex derived from theoretical analysis. 3) Determine the distinct mechanisms for targeting hydrolysis at a specific site. Endonucleases Fokl, Sill and Bsll recognize and cleave DNA by mechanisms that differ from most restriction enzymes. We will determine the structure of Bsll, which is unusual in its heterotetrameric architecture and has a clinical application in detecting cancerous mutations. We will complete the structure of Sill and cocrystallize the enzyme with a set of non-cognate and varied cognate DNA sites. Finally, we will complete our analysis of Fokl and cocrystallize it in an activated synaptic form with two DNA molecules. | |||||||
| > > | Abstract: DESCRIPTION (provided by applicant): Protein-DNA selectivity is a central event in many biological processes, ranging from transcription and replication to restriction and modification. Type II restriction endonucleases are ideal systems for studying selectivity because of their high specificity and great variety. The enzymes recognize and cleave DNA sequences that vary between four to eight base pairs. Their sequence specificity is remarkable. A single base pair change within the recognition sequence can lead to well over a million fold reduction in activity. An understanding of sequence specific cleavage is relevant to protein mediating site-specific recombination and DNA repair by excision. The long-term goals of this project are to understand the mechanism by which type II restriction enzymes recognize and cleave DNA, and to design mutants with altered specificities. The three broad aims are: 1) Determine the basis of discrimination between closely related DNA sites. With structures of BamHI? and Bglll in hand, we will now determine structures of an "intermediate" endonuclease with the specificity of both BamHI? and Bglll: BstYI. We will also manipulate the specificity of BamHI? with the aid of these new structures. 2) Determine the basis of specific versus non-specific DNA binding. We have determined the structure of BamHI? bound to one non-cognate DNA sequence. We will now determine structures of BamHI? bound to other non-cognate DNA sequences, in order to see the structural adaptations in going from one non-cognate sequence to another. We will also experimentally test a model of the non-cognate complex derived from theoretical analysis. 3) Determine the distinct mechanisms for targeting hydrolysis at a specific site. Endonucleases Fokl, Sill and Bsll recognize and cleave DNA by mechanisms that differ from most restriction enzymes. We will determine the structure of Bsll, which is unusual in its heterotetrameric architecture and has a clinical application in detecting cancerous mutations. We will complete the structure of Sill and cocrystallize the enzyme with a set of non-cognate and varied cognate DNA sites. Finally, we will complete our analysis of Fokl and cocrystallize it in an activated synaptic form with two DNA molecules. | |||||||
| Thesaurus Terms: DNA, chemical cleavage, enzyme mechanism, enzyme structure, restriction endonuclease DNA binding protein, active site, enzyme activity, enzyme substrate complex, hybrid enzyme, hydrolysis, nucleic acid sequence, structural biology X ray crystallography, crystallization, fluorescence resonance energy transfer Institution: MOUNT SINAI SCHOOL OF MEDICINE OF NYU OF NEW YORK UNIVERSITY NEW YORK, NY 10029 Fiscal Year: 2005 Department: PHYSIOLOGY AND BIOPHYSICS Project Start: 01-APR-1990 Project End: 31-MAR-2008 ICD: NATIONAL INSTITUTE OF GENERAL MEDICAL SCIENCES IRG: ZRG1 | ||||||||
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