The Role of Zinc in the Interaction of TAX1BP1 and Itch Studied by Fluorescence Anisotropy and Structural Modeling Ashley F. Cowan, Joel E.P. Brandis,

Slides:

Advertisements

Similar presentations

Understanding biology through structures Course work 2006 Protein-Nucleic Acid Interactions: General Principles.

Advertisements

9-1 Chapter 9 DNA-Protein Interactions in Bacteria Student learning outcomes: Describe examples of structure /function relationships in phage repressors.

Synthetic approaches to transcription factor regulation and function Tim Johnstone BIOL1220 Spring 2010.

Dual conformations for the HIV-1 gp120 V3 loop in complexes with different neutralizing Fabs RL Stanfield, E Cabezas, AC Satterthwait, EA Stura, AT Profy,

CHAPTER 8 Metabolic Respiration Overview of Regulation Most genes encode proteins, and most proteins are enzymes. The expression of such a gene can be.

1 Genetic regulation Genotype is not phenotype: bacteria possess many genes that they are not using at any particular time. Transcription and translation.

Group 1 Biology - Chemistry Interface course make up: introductory course emphasizes integration of chemical & biological concepts life science majors.

Objectives for protein section II Explain conformational changes in proteins. Explain how GTP binding & hydrolysis allows ras & other G-proteins to function.

Protein Structure Lecture 2/26/2003. beta sheets are twisted Parallel sheets are less twisted than antiparallel and are always buried. In contrast, antiparallel.

Mapping of Calmodulin Binding Sites on the IP 3 R1 N. Nadif Kasri, I. Sienaert, J.B. Parys, G. Callewaert, L. Missiaen and H. De Smedt Laboratory of Physiology,

Protein-protein interactions between widely-expressed and testis specific subunits of TFIIA and TFIID in Drosophila melanogaster Leah Hirschman, Mark Hiller.

Introduction to Protein Structure

Comparison of p53 Structure: Wild type vs. mutant What change in wild type p53 may lead to cancer?

P53 Missense Mutation Cancer. Outline Disease related to p53 Role and regulation pathway Structure of p53 Missense mutation and consequences Experiment’s.

 I can describe the structure and explain the significance and functions of enzymes in biological systems › I can describe why an investment of activation.

Trp Operon A brief description. Introduction a repressible system In this system, though, unlike the lac operon, the gene for the repressor is not adjacent.

The importance of telomerase in maintaining chromosome integrity.

Higher Biology Chapter 16 Gene Mutations. This type of mutation involves a change in one or more of the nucleotides in a strand of DNA. There are four.

Chapter 9 From DNA to Protein.

Changes In Protein Sequences Of the HIV-1 gp120 V3 Region In Non-Progressor Types Nicki S.Harmon Samantha M. Hurndon LMU Department of Biology BIOL 368.

KIMBERLY JONES AUGUST 20, 2010 SPUR PROGRAM 2010 UNIVERSITY OF OREGON BERGLUND LAB INSTITUTE OF MOLECULAR BIOLOGY P.I.-ANDY BERGLUND, PHD MENTORS-AMY MAHADY,

Specific Defenses of the Host Part 2 (acquired or adaptive immunity)

Fanconi Anemia Anthony Winchell. What is Fanconi Anemia?

V3 Loop Binding to CCR5 and CXCR4 of Rapid and Non Progressor HIV Patients in Baltimore MD BackgroundCCR5 and CXCR4 Receptor Rapid-progressor Patient 10.

Haemoglobin: Structure and Formation of Proteins 1.1.2, a, b,c.

Date of download: 9/18/2016 Copyright © 2016 American Medical Association. All rights reserved. From: Molecular Basis of Low-Penetrance Retinoblastoma.

19.5 Protein Structure: Tertiary and Quaternary Levels

Alignment table: group 4

                                  

“Simulations of the dynamics at an RNA-protein interface”

Figure 3. Kinetics effects of the C domain on ER–EREc binding

Figure 1 Dependence of DAPI displacement on RecA protein concentration

Regulation of Gene Expression in Eukaryotes

CHEMISTRY 2 BIOCHEMISTRY.

Superantigen Recognition by γδ T Cells

Glen S. Cho, Jack W. Szostak Chemistry & Biology

Engineering a Protein Scaffold from a PHD Finger

Beyond the “Recognition Code”

LMU Department of Biology

Volume 115, Issue 2, Pages (October 2003)

Dual conformations for the HIV-1 gp120 V3 loop in complexes with different neutralizing Fabs RL Stanfield, E Cabezas, AC Satterthwait, EA Stura, AT Profy,

Structural Basis for the Specific Recognition of Methylated Histone H3 Lysine 4 by the WD-40 Protein WDR5 Zhifu Han, Lan Guo, Huayi Wang, Yue Shen, Xing.

Volume 91, Issue 6, Pages (December 1997)

Hung-Ta Chen, Steven Hahn Cell

Scot A Wolfe, Elizabeth I Ramm, Carl O Pabo Structure

Volume 25, Issue 5, Pages e3 (May 2017)

A Shared Surface of TBP Directs RNA Polymerase II and III Transcription via Association with Different TFIIB Family Members Xuemei Zhao, Laura Schramm,

Rachel L Winston, Joel M Gottesfeld Chemistry & Biology

Volume 23, Issue 5, Pages (May 2015)

Volume 87, Issue 2, Pages (October 1996)

Volume 18, Issue 6, Pages (June 2010)

Structure, Exchange Determinants, and Family-Wide Rab Specificity of the Tandem Helical Bundle and Vps9 Domains of Rabex-5 Anna Delprato, Eric Merithew,

Volume 11, Issue 2, Pages (February 2003)

A Role for Intersubunit Interactions in Maintaining SAGA Deubiquitinating Module Structure and Activity Nadine L. Samara, Alison E. Ringel, Cynthia Wolberger

Mechanisms Contributing to T Cell Receptor Signaling and Assembly Revealed by the Solution Structure of an Ectodomain Fragment of the CD3ϵγ Heterodimer

Volume 14, Issue 4, Pages (April 2006)

Structure of the Staphylococcus aureus AgrA LytTR Domain Bound to DNA Reveals a Beta Fold with an Unusual Mode of Binding David J. Sidote, Christopher.

Protein Folding and Unfolding at Atomic Resolution

Volume 11, Issue 1, Pages 1-12 (April 2015)

Volume 2, Issue 1, Pages 1-4 (January 1994)

Amino acid sequence deduced from the sequence of contig 131 of G

Volume 113, Issue 9, Pages (November 2017)

Volume 18, Issue 3, Pages (March 2011)

Mark S. Dunstan, Debraj GuhaThakurta, David. E. Draper, Graeme L. Conn

Volume 102, Issue 1, Pages (January 2012)

Guilty as charged Cancer Cell

Volume 7, Issue 1, Pages 1-11 (July 1997)

Just the Beginning: Novel Functions for Angiotensin-Converting Enzymes

Structure of GABARAP in Two Conformations

Presentation transcript:

The Role of Zinc in the Interaction of TAX1BP1 and Itch Studied by Fluorescence Anisotropy and Structural Modeling Ashley F. Cowan, Joel E.P. Brandis, and Barbara T. Amann Department of Biology and Chemistry, Goucher College, 1021 Dulaney Valley Rd., Baltimore, MD Abstract Zinc deficiency leads to a suppressed immune system although the mechanism is not fully understood. The Nuclear Factor kappa B (NF-kB) signaling pathway turns inflammation on and off. Many key proteins in this pathway bind to zinc. The tightest interaction occurs between of the second TAX1BP1 zinc finger (TAXF2) and the first Itch WW domain (WW1) domain when zinc is present. Through mutational studies, we have determined that four amino acids residues located in the TAXF2 are responsible for this increase affinity in the presence of zinc. Background Zinc deficiency affects 2 billion people worldwide and leads to major health problems including a suppressed immune system. The NFkB signaling pathway has many zinc binding proteins, which may explain why zinc leads to a suppressed immune system. Zinc binding proteins are necessary for turning on and off inflammation. Within the NFkB pathway, the A20 complex shuts off inflammation through negative feedback loop. TAX1BP1 and Itch are integral parts of the A20 complex. The TAX1BP1 and Itch proteins interact with one another through two PPXY zinc finger domains and four WW domains, respectively. Results Conclusion Of the TAXF1 mutants, TAXF1B2 binds the strongest to WWI and has the most similar K d value to TAXF2. TAXF1KKCPLCELMFPPNYDQSKFEEHVESHFK TAXF1b2KKCPMCEEQFPPNYDQQKFERHVQTHFD TAXF2KVCPMCSEQFPPDYDQQVFERHVQTHFD Modeling WW domain is a small three beta strand structure found in many proteins. Each TAX1BP1 zinc finger contains a PPXY domain and binds zinc with two conserved cysteines and histidines. TAX1BP1 fingers were modeled using a classical zinc finger. This causes the PPXY sequence to curve. Goal To determine what amino acids in TAXF2 allows it to bind the strongest to WWI when zinc is present. Methods Mutations were made to specific amino acids to turn TAXF1 into TAXF2: TAXF1KKCPLCELMFPPNYDQSKFEEHVESHFK TAXF1aKKCPMCELQFPPNYDQQKFEEHVETHFK TAXF1b1KVCPMCSLQFPPDYDQQVFEEHVETHFK TAXF1b2KKCPMCEEQFPPNYDQQKFERHVQTHFD TAXF2KVCPMCSEQFPPDYDQQVFERHVQTHFD Experimental Procedure CC1CC2CC3 PPXY ZF1 PPXY ZF2 C2 HECTWW1WW2WW3WW4 TAX1BP1 ITCH RNF11 A20 A20 Complex includes the proteins RNF11, Itch, TAX1BP1, and A20 WW domainClassic TFIIA zinc finger and PPXY region unattached Model of TAXF2 zinc finger with PPXY region in red Synthetic peptides purified by HPLC. Binding activity of cobalt to protein quantified by uv/vis spectroscopy. Cobalt then substituted by zinc. K d values of zinc fingers bond to WWI determined from fluorescence anisotropy. Figure 1. Anisotropy compared to the concentrations of TAXF2 or mutants of TAXF2 when added to WWI. Graph produced through the use of KaleidaGraph. The amino acids in red are those determined to be important to the interaction of TAXF2 and WWI. Future experiments could lead to us discovering the specific amino acid(s) responsible for the increased bonding. Table 1. K d values in  M of TAX1BP1 domains TAXF2 and TAXF1 with mutations bond to Itch WW1. All TAX1BP1 domains contain zinc. K d values are averaged from 2-3 different trials. + Zinc