Walter G. Gonzalez

In this site you will find information on projects and interests relating to my work in the lab of Dr. Jaroslava Miksovska. Through the years I have become interested in the use of theorethical and experimental techniques to elucidate nature's intricate mechanisms both at the cellular and molecular level. By combining a wide range of techniques I have been able to reveal the intricate mechanism of action of small ligands on calcium binding proteins as well as characterizing numerous fluorophores.

Curriculum Vitae

Events

  • Will be attending the 10th LFD Workshop in Advanced Fluorescence Imaging and Dynamics.October 26th to 30th, UC Irvine, CA
  • Attended ACS Boston conference to network/career development and to find the latest in chemistry research. August 13th to August 23rd, 2015.
  • Presented Network/Allostery relationship on DREAM at FAME 2015 conference. May 7th to 9th, 2015 at Tampa, FL

Background

I am currently a fifth year PhD candidate at Florida International University working under supervision of Dr. Jaroslava Miksovska. I specialize in spectroscopic techniques to study the biophysical properties of proteins in-vitro. I also implement calorimetric techniques in order to study optically silent processes ranging from nansoeconds to minutes. Furthermore, through the use of molecular dynamics, docking and quantum mechanics I am currently gainign insight into the role of protein dynamics in allosteric control as well as drug binding. Altogether, the combination of these techniques provide unparallel ability to study structure-function relationship in proteins and how ligand binding can be used to modulate their activity. Understanding of these mechanisms allow me and our to propose approaches to regulate biological processes such as neuronal and cardiac signaling as well as to elucidate the pathology of diseases such as a Alzheimer's disease, arrythmias and hypertension.

  • Absorption spectroscopy
  • Fluorescence spectroscopy
  • Frequency domain lifetime
  • Time domain luminiscence New
  • Polarization
  • Frequency domain anisotropy
  • Photothermal beam deflection
  • Photoacoustic calorimetry
  • Isothermal calorimetry New
  • Molecular Dynamics New

Biology

In the lab of Dr. Jaroslava Miksovska at FIU we employ the use of cutting edge bioengineering in order to study the effect of mutations, deletions and more radical changes on protein actvity, structure, and dynamics. Some of these techniques include:

  • PCR
  • Sequencing
  • Cloning
  • Large scale protein production
  • Site-directed mutagenesis
  • Cell screening

Physics

My initial training in physics gives me an unique perspective into the features of protein strucutre. With support from the MBRS program at FIU I have independently built two GPU-CPU workstations and developed a robust routine for conducting ultrafast molecular dynamic simulations using the vmd-namd package. These simulations provide detailed insight into the role of specific amino acids in protein structure and dynamics.

  • Molecular Dynamics NAMD/GROMACS
  • Docking
  • Ligand parameterization
  • Quantum mechanics
  • Density functional theory
  • Visualization VMD

Chemistry

As part of the Chemistry and Biochemistry department I have been mainly focused in the chemistry of proteins s well as protein ligands. Specifically, I enjoy understanding the chemistry of drugs known to bind to and modulate the activity of calcium binding proteins and to exploit these chemical features to elucidate the mechanism of action. I am also particularly interested in the chemistry of fluorescent molecules and the underlying photophysics that arise from specific chemical groups.

  • Enzyme reactions
  • Protein post-translational modifications
  • Drug discovery
  • Fluorophore design
  • Graphene Oxide
  • Unnatural amino acids
13

Completed Projects

90

Citing articles

11

Awards Achievements

18

Conference Presentations

Experience

My current PhD work involves the use of a wide array of theoretical and experimental biophysics to study the mechanism of signal transduction in calcium binding proteins, with special emphasis on the neuronal calcium sensor superfamily and their role in voltage-gated potassium current regulation. More specifically, much of my research involves investigating the role of specific amino acid(s) on the activity of a protein, first by conducting various in-silico theoretical simulations followed by extensive characterization of protein mutations using biochemical techniques in-vitro.

Techniques

Current skills and expertise.

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Absorption spectroscopy

Absorption spectroscopy is an essential technique that allow us to accurately determine sample concentrations as well as easily access any ground state changes in a wide range of chromophores.

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Fluorescence spectroscopy

This techique is widely applied in science and I currently focus on using it to investigate the excited state environments of extrinsic and intrinsic fluorophores in proteins as well other fluorescent molecules.

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Depolarization

Fluorescence depolarization is a robust technique that allow us to measure the rotational diffusion of small and large molecules in solutions as well as intermolecular distances. Steady state and time resolved depolarization (anisotropy) also allows the fascile determination of binding events and oligomerization states.

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Photothermal

We can also employ photothermal techniques such as PBD (phototherml beam deflection) and PAC (photoacoustic calorimetry) in order to observe the kinetics of processes which are optically silent. Additionally, these techniques permit the measurement of processes ranging from nanoseconds to milliseconds. Both PBD and PAC provide themodynamics parameters such as the activation energy, change in volume and change in enthalpy for a specific process.

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Molecular Dynamics

By analysing the molecular motion of atoms in proteins using the molecular dynamics simulation package VMD and NAMD we can gain detailed informaiton on at the atomic level. This theorethical approach provide us with a greater insight into the role of specific amino acids/domains and network of amino acids in proteins.

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Bioengineering

By applying PCR, cloning, and site directed mutagenesis we are able to express and purifiy a wide variety of proteins in large quantities, which is an indespensable step for biochemical characterization. Site directed mutagenesis also allow us to study the structure-function relationship of proteins.

Publications

Recent projects.

Amphiphilic residues 29-44 of DREAM N-termini mediate the calmodulin:DREAM complex formation.

Biochemistry, (2015)54:28:4391–4403

Here we identify that amino acids at the N-terminus of DREAM (residues 29-44) modulate the association with calmodulin. We show that CaM stabilizes expression of DREAM and can modulate DREAM:DNA interactions.

Strain Promoted Click Chemistry Nucleosides and Application to Living Cell Fluorescent Imaging.

Bioconjug Chem. (2015)in press

In collaboration with Dr. Wnuk and Dr. Zayas we characterized the photophysical properties of novel fluroescence click nucleosides.

Modulation of conformational dynamics and stability of DREAM

Protein Sci. (2015)5:741-51

In this study Khoa Pham in collaboration with Dr. Chapagain show that metal binding to DREAM can modulate the local conformational dynamics as well as stability.

Modulation of Kv4.3:KChIP3 interactions by NS5806.

J. Biol. Chem. (2014)289:32201-32213

Here we show that the novel Ito current activator NS5806 bind at the C-terminus of KChIP3 (DREAM) and enhances the binding to the N-terminus of the Kv4 potassium channels.

Site-Specific Dynamics of Aβ1-23 Amyloid Formation

Chem. Commun. (2015),51:32:7000-3

The dynamics of Aβ1-23 amyloid formation are revealed by a wide variety of techniques including atomic force miscroscopy, IR spectrsocopy, and steady state and time resolved fluroescence/anisotropy.

Flavin Adenine Dinucleotide Structural Motifs: From Solution to Gas Phase

Anal. Chem. (2014),86:20:10223–30

Dr. Fernandez-Lima abd coworkers here show that flavin adenine can populate multiple conformations and they are shown it all their complexity by high resolution TIMS-MS, quantum mechanics and fluroescence.

Full publication list

Funding

Thanks to the following funding for supporting my research.

MBRS RISE fellowship



Summer 2015 to present


NIH/NIGMS R25 GM061347


website

BRI Summer Research Award



Summer 2013, 2014 and 2015


NIH/NIGMS R25 GM061347


Website

National Science Foundation



RA support/supplies 2011-2014


MCB 1021831 to Dr. Miksovska


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Contact Feel free to contact me

Dr. Jaroslava Miksovska's Lab
Florida International University
11200 SW 8th Street Owa Ehan 320
Miami, Florida 33199
Office: (305) 348-4262
Walter G. Gonzalez
wgonz002@gmail.com