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Title: Protein expression, characterization and activity comparisons of wild type and mutant DUSP5 proteins

Journal Article · · BMC Biochemistry (Online)
 [1];  [1];  [2];  [1];  [3];  [3];  [1];  [1];  [4];  [3];  [2];  [5]
  1. Medical College of Wisconsin, Milwaukee, WI (United States)
  2. Marquette Univ., Milwaukee, WI (United States)
  3. Concordia Univ. of Wisconsin, Mequon, WI (United States)
  4. Argonne National Lab. (ANL), Argonne, IL (United States)
  5. Medical College of Wisconsin, Milwaukee, WI (United States); CRI Developmental Vascular Biology Program, Milwaukee, WI (United States)
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Background: The mitogen-activated protein kinases (MAPKs) pathway is critical for cellular signaling, and proteins such as phosphatases that regulate this pathway are important for normal tissue development. Based on our previous work on dual specificity phosphatase-5 (DUSP5), and its role in embryonic vascular development and disease, we hypothesized that mutations in DUSP5 will affect its function. Results: In this study, we tested this hypothesis by generating full-length glutathione-S-transferase-tagged DUSP5 and serine 147 proline mutant (S147P) proteins from bacteria. Light scattering analysis, circular dichroism, enzymatic assays and molecular modeling approaches have been performed to extensively characterize the protein form and function. We demonstrate that both proteins are active and, interestingly, the S147P protein is hypoactive as compared to the DUSP5 WT protein in two distinct biochemical substrate assays. Furthermore, due to the novel positioning of the S147P mutation, we utilize computational modeling to reconstruct full-length DUSP5 and S147P to predict a possible mechanism for the reduced activity of S147P. Conclusion: Taken together, this is the first evidence of the generation and characterization of an active, full-length, mutant DUSP5 protein which will facilitate future structure-function and drug development-based studies.

Research Organization:
Argonne National Laboratory (ANL), Argonne, IL (United States)
Sponsoring Organization:
USDOE
Grant/Contract Number:
AC02-06CH11357
OSTI ID:
1201767
Journal Information:
BMC Biochemistry (Online), Vol. 15, Issue 1; ISSN 1471-2091
Publisher:
BioMed CentralCopyright Statement
Country of Publication:
United States
Language:
English
Citation Metrics:
Cited by: 10 works
Citation information provided by
Web of Science

References (39)

ZDOCK server: interactive docking prediction of protein-protein complexes and symmetric multimers journal February 2014
Evaluation of secondary structure of proteins from UV circular dichroism spectra using an unsupervised learning neural network journal January 1993
Regulation of the inducible nuclear dual-specificity phosphatase DUSP5 by ERK MAPK journal December 2009
ERK1/2 MAP kinases: Structure, function, and regulation journal August 2012
DICHROWEB: an interactive website for the analysis of protein secondary structure from circular dichroism spectra journal January 2002
Mechanism of Cdc25B Phosphatase with the Small Molecule Substrate p -Nitrophenyl Phosphate from QM/MM-MFEP Calculations journal April 2009
Vascular tumors of infancy and childhood: beyond capillary hemangioma journal November 2006
Crystal structures of fusion proteins with large-affinity tags journal July 2003
Experimental Validation of the Docking Orientation of Cdc25 with Its Cdk2−CycA Protein Substrate † journal December 2005
Dual-specificity MAP kinase phosphatases (MKPs) and cancer journal March 2008
Crystal structure of the catalytic domain of human DUSP5, a dual specificity MAP kinase protein phosphatase journal October 2006
Dusp-5 and Snrk-1 coordinately function during vascular development and disease journal January 2009
Comparison of simple potential functions for simulating liquid water journal July 1983
Structural Mechanism of Oxidative Regulation of the Phosphatase Cdc25B via an Intramolecular Disulfide Bond † , ‡ journal April 2005
Flipped Out:  Structure-Guided Design of Selective Pyrazolylpyrrole ERK Inhibitors ‡ journal March 2007
Increasing the precision of comparative models with YASARA NOVA-a self-parameterizing force field journal May 2002
Dual-specificity MAP kinase phosphatases (MKPs): Shaping the outcome of MAP kinase signalling journal August 2012
Making optimal use of empirical energy functions: Force-field parameterization in crystal space journal January 2004
ConSurf 2010: calculating evolutionary conservation in sequence and structure of proteins and nucleic acids journal May 2010
Improving physical realism, stereochemistry, and side-chain accuracy in homology modeling: Four approaches that performed well in CASP8 journal January 2009
Catalytic Activation of the Phosphatase MKP-3 by ERK2 Mitogen-Activated Protein Kinase journal May 1998
Signal-dependent repression of DUSP5 by class I HDACs controls nuclear ERK activity and cardiomyocyte hypertrophy journal May 2013
ERK Signaling Is a Molecular Switch Integrating Opposing Inputs from B Cell Receptor and T Cell Cytokines to Control TLR4-Driven Plasma Cell Differentiation journal October 2006
PROSESS: a protein structure evaluation suite and server journal May 2010
A smooth particle mesh Ewald method journal November 1995
Pathogenesis of Vascular Anomalies journal January 2011
Solution Structure of the MAPK Phosphatase PAC-1 Catalytic Domain journal February 2003
A point-charge force field for molecular mechanics simulations of proteins based on condensed-phase quantum mechanical calculations journal October 2003
Structure and regulation of MAPK phosphatases journal July 2004
Mechanistic Basis for Catalytic Activation of Mitogen-activated Protein Kinase Phosphatase 3 by Extracellular Signal-regulated Kinase journal March 2000
In Vitro Enzymatic Assays of Protein Tyrosine Phosphatase 1B journal May 2001
Activation Mechanism of the MAP Kinase ERK2 by Dual Phosphorylation journal September 1997
Solution Structure of ERK2 Binding Domain of MAPK Phosphatase MKP-3 journal February 2001
Structural Mechanism of Oxidative Regulation of the Phosphatase Cdc25B via an Intramolecular Disulfide Bond journal April 2005
T-cell Development and Function Are Modulated by Dual Specificity Phosphatase DUSP5 journal April 2008
Spatiotemporal Regulation of ERK2 by Dual Specificity Phosphatases journal July 2008
Inhibition of Mitogen-activated Protein Kinase Phosphatase 3 Activity by Interdomain Binding journal August 2008
Specific Inactivation and Nuclear Anchoring of Extracellular Signal-Regulated Kinase 2 by the Inducible Dual-Specificity Protein Phosphatase DUSP5 journal February 2005
ERKs in Cancer: Friends or Foes? journal January 2014

Cited By (3)

Identification of inhibitors that target dual-specificity phosphatase 5 provide new insights into the binding requirements for the two phosphate pockets journal August 2015
Serendipitous discovery of light-induced (In Situ) formation of an Azo-bridged dimeric sulfonated naphthol as a potent PTP1B inhibitor journal May 2017
Extracellular signal-regulated kinases 1/2 as regulators of cardiac hypertrophy journal July 2015

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Related Subjects

59 BASIC BIOLOGICAL SCIENCES
DUSP5
mutation
vascular anomalies
protein purification
molecular modeling