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Rajagopal Appavu, Ph.D.
Department of Pharmacology and Toxicology
Sealy Center for Vaccine Development
University of Texas Medical Branch-Galveston, Texas
E-mail: [email protected]
[email protected]
Mobile: (409) 256-9987
SKILLS SUMMARY
• Strong background in the pre-clinical vaccine development of protein/peptide (nanomaterial) based vaccine adjuvant
design, synthesis (Malaria vaccine, Melanoma vaccine, T-Cell vaccine, Cocaine vaccine, and virus-like particles etc.).
Protein/peptide adjuvants chemical modification, peptidomimetics, fluorescence labeling, peptide-nucleic acid
conjugation, peptide-small molecule coupling, purification, structural characterization of polypeptides (NMR, TEM, Xray, FT-IR, CD, Mass Spectrometry [MALDI TOF/TOF, LC-MS, GC-MS], DSC, TGA, HPLC, and DTA).
• Excellent skills in Immunology using flow cytometry, ELISA, animal vaccination/immunization and behavior studies,
blood serum isolation, B/T Cell studies, Cytotoxicity assays, and examining antibody responses in animal model system
(humoral immune response). Working experience upto ABSL2. Live cell imaging in mice (CT, MRI, and PET).
• Technical expertise in mammalian cell culture, hybridoma selection, native and SDS PAGE, Western analysis,
protein purification, immunostaining & fluorescence microscopy, and immunoprecipitation.
• Expert in alliance management both in academic and industrial collaboration.
• Excellent software skills in Flowjo, Insight, PyMOL, MOLMOL, ORIGIN, ChemDraw, Flowjo, and NMR-Topspin.
• Strong academic communication skills – oral and written (grant writing, publications, and teaching).
EMPLOYMENT
Postdoctoral research scholar, Uni. of Texas Medical Branch-Galveston, USA
Mar 2012-till date
Senior Research Fellow, Molecular Biophysics Unit, IISc-Bangalore, India
Aug 2007-Feb 2011
Junior Research Fellow, Molecular Biophysics Unit, IISc-Bangalore
Aug 2005-Jul 2007
Research Assistant, Molecular Biophysics Unit, IISc-Bangalore
Mar 2005-Jul 2005
EDUCATION
Ph.D. in Molecular Biophysics (Biochemistry)
2012
Thesis Title: Designed Synthetic Peptides: Models for Studies of Conformation Transitions and Aromatic Interactions
Indian Institute of Science-Bangalore, India
M.Sc. in Chemical Sciences
2004
Pondicherry University, Puducherry
Thesis title: Cyclic Voltammetric studies on Synthetic Cobalt Complexes
B.Sc. in Chemistry
2001
Govt. Arts College, Salem, Tamil Nadu
PUBLICATIONS
ŸAppavu Rajagopal, Chesson, C. Brent, Sara M
Dann, and Jai S Rudra, Enantiomers of self-assembling peptides elicit
inverse antibody and CD8 T cell responses, Biomaterials (Manuscript in Preparation).
ŸAppavu Rajagopal, Rajkishor Rai, Srinivasarao Raghothama, Narayanaswamy Shamala, and Padmanabhan Balaram,
Design of a Peptide Hairpin Containing a Central Three-Residue Loop, (Manuscript in Preparation).
ŸAppavu
Rajagopal, Subrayashastry Aravinda, Srinivasarao Raghothama, Narayanaswamy Shamala, and Padmanabhan
Balaram, Chain Length Effects on Helix-Hairpin Distribution in Short Peptides with Aib-DAla and Aib-Aib Segments,
Biopolymers (PeptideScience), DOI:10.1002/bip.21613.
ŸRaghavender
Upadhyayula, Chatterjee Bhaswati, Saha Indranil, Appavu Rajagopal, Narayanaswamy Shamala, and
Padmanabhan Balaram, Entrapment of a Water Wire in a Hydrophobic Peptide Channel with an Aromatic Lining, The
Journal of Physical Chemistry B, DOI: 10.1021/jp200462h.
Ÿβ-turn
analogs in model αβ hybrid peptides. Structural characterization of peptides containing β2,2Ac6c and β3,3Ac6c
residues. Krishnayan Basuroy, Appavu Rajagopal, Srinivasarao Raghothama, Narayanaswamy Shamala, Padmanabhan
Balaram, Chemistry-An Asian Journal, DOI: 10.1002/asia.201200052.
ŸAppavu
Rajagopal, Subrayashastry Aravinda, Srinivasarao Raghothama, Narayanaswamy Shamala, and Padmanabhan
Balaram, Aromatic Interactions in Model Peptide β-Hairpins: Ring Current Effects on Proton Chemical Shifts
Biopolymers (PeptideScience), DOI: 10.1002/bip.22003.
COURSE/CONFERENCE/WORKSHOPS
• EMBO Global Exchange Lecture Course on Molecular Mechanism of Protein Transport
Dec 1-8, 2010
• International Symposium on Interface between Chemistry and Biology
Feb 17-19, 2009
• Conference on Nuclear Magnetic Resonance Society-India
Feb 2-5, 2009
• IISc Centenary Conference on Biology and Pathogenesis of Viruses: Molecular Insights
May 4-5, 2009
• Conference on 5th Junior National Organic Symposium Trust-India
Dec 4-7, 2009
• Bruker Conference & Workshop on Mass Spectrometry
Dec 4-7, 2008
• “Sushi and the Snail: A meeting on cellular trafficking, circuits and behaviour”
Feb 23-24, 2007
• NCBS-JNCASR-Harvard Symposium on “Interdisciplinary Materials Science Computational & Biology”
Aug, 2006
COURSES AT RESEARCH PROGRAMME
Introduction to Biophysical Chemistry, Molecular Spectroscopy and Structure, Conformational and Structural Aspects of
Biopolymers, Protein Structure: Folding and Design, Biological Chemistry: Molecular Approach, and Introduction & Biomolecular Applications of 2D NMR Spectroscopy
AWARDS
Wellcome Trust, UK (Travel award for International Conference on Biomolecular Forms and Functions)Jan 2013
Senior Research Fellowship, Department of Science and Technology, Govt. of India, New Delhi
Junior Research Fellowship, Molecular Biophysics Unit, IISc-Bangalore
National Entrance Test for Research Admission, Indian Institute of Science-Bangalore
Graduate Aptitude Test in Engineering (GATE) - MHRD, Govt. of India, New Delhi
Aug 2010-Jul 2013
Aug 2005- Jul 2010
Awarded 2005
Awarded 2004
PRESENTATIONS
ORAL: Design of peptide models for β-hairpins and equilibrating helix-hairpin structures
POSTER: NMR analysis of solvent dependent conformational transitions in designed octapeptides
Dec 4-7, 2009
Feb 2-5, 2009
RESEARCH INTEREST
• Broad curiosity in development of peptide/proteins as vaccine candidates, which includes protein-protein interaction
analysis, synthesis of small molecules (organic/ inorganic/natural products), peptides (cyclic/acyclic disulfide bond
formation), synthetic proteins via native chemical ligation approach, solution and solid-phase peptide synthesis and
introduction of diverse chemical bonds in synthetic proteins
• Structural determination of protein-DNA (Solid: X-ray, Solution: 2D & 3D NMR, and Gas: Mass Spectrometry)
interactions and their pathways, stability, dynamics, and function
• Molecular modeling and structure based drug design. Design, synthesis, and development of MRI contrast agents.
• Molecular mechanism of intracellular traffic (endocytosis and exocytosis) incorporation ligand free-bound. Further,
biophysics and biochemistry of nanoparticle peptide disease targets.
CURRENT RESEARCH
1. Enantiomers of self-assembling peptides elicit inverse antibody and CD8 T cell responses
Self-assembling fibrillar peptide biomaterials have been investigated for various biotechnological applications and been
developed based on several different secondary structures like α-helices, β-sheets, β-hairpins, and peptide amphiphiles.
However, the bulk of the research has been focused on β-sheet fibrillizing peptides. Several different peptides forming βsheet fibrils have been studied, including strictly alternating polar/nonpolar peptides, peptides from extracellular matrix
proteins, designed glutamine-rich peptides, and sequences from amyloid-forming proteins. Recently it was reported that the
glutamine-rich self-assembling peptide Q11 (Ac-QQKFQFQFEQQ-Am) acts as an immune adjuvant when conjugated to
peptide antigens. Fusion peptides of Q11 and an antigenic peptide OVA323-339 (H2N-ISQAVHAAHAEINEAGR-OH from
chicken egg ovalbumin) raised strong and long-lived anti-OVA323-339 antibody responses in mice. Antibodies produced by
Q11 nanofibers conjugated to peptide epitopes have been shown to be protective in mouse models of malaria and cancer.
Another self-assembling amphipathic peptide KFE8 (Ac-KFFEFKFE-Am) was also reported to behave similar to Q11
when conjugated to OVA-OVA323-339. To investigate the nature of the amino acid sequence on immune responses, we
synthesized the self-assembling peptide KFE8 and its enantiomeric version (D)KFE8 composed of D-amino acids and
coupled them to OVA323-339. Mice were immunized with nanofibers of OVA323-339-(L)KFE8 or OVA323-339-(D)KFE8 and
antibody responses were investigated.
2. Virus Inspired Self-assembling Peptide Nanoparticle Vaccines
Nanoparticles constructed from self-assembly of plant and animal viral capsid proteins have found numerous applications in
biotechnology. However, synthesis of modified viral coat proteins requires gene recombination and protein expression,
which are complicated and susceptible to contamination. Alternatively, the reconstruction of virus-like nanoparticles from
synthetic peptides would enhance their potential for in vivo applications and virus-inspired C3- and C5-symmetric selfassembling peptide nanoparticles have been investigated for encapsulation of small guest molecules including drugs and
DNA. Here we tested whether virus-inspired self-assembling peptides could be used as antigen delivery scaffolds in vaccine
development and immunotherapy. We synthesized a 24-mer self-assembling peptide (SAP) from the tomato bushy stunt
virus (TBSV) that has been reported to assemble into C3-symmetric nano particles with the C-terminus directed towards the
particle exterior. We hypothesized that appending an antigenic CD8 T cell epitope OVA (chicken ovalbumin aa 257-264);
to the C terminus of the SAP via a short linker would result in nanoparticles that display OVA on the surface (SAP-OVA).
Based on our previous work with self-assembling peptide nanofibers, we expected that C3-symmetric SAP-OVA
nanoparticles would elicit robust CD8 T cell responses without the need for any exogenous adjuvants.
3. Synthetic Nanofiber Vaccines for Cocaine Addiction
Cocaine addiction is second only to opiates as the most problematic drug in the Americas with an enormous fee in human
tragedy as well as in public health and safety. Currently there is no FDA approved pharmacological therapy for cocaine
addiction, which has prompted the development of immuno-prophylactic alternatives. A therapeutic vaccine that elicits
anti-cocaine antibodies will allow the rapid sequestration of the drug in circulation and reduce the amount and rate of its
entry into the brain. A limiting factor to the success of small molecule drug vaccines is the low degree of immunity evoked
by the addictive drug analog and lack of effective yet safe immune adjuvants. The goals of this proposal are to design and
develop cocaine vaccines based on designed cocaine analogs in combination with a peptide nanofiber-based delivery
platform for eliciting high titers of anti-cocaine antibodies and test their efficacy in a mouse motor activity assay. The work
is divided into two aims: Aim 1) Design and synthesis of self-assembling peptide nanofiber-based cocaine vaccines. Aim 2)
Test the efficacy of peptide nanofiber cocaine vaccine formulations in a mouse model. Our approach will be to design and
chemically synthesize peptide nanofiber vaccines against cocaine. In aim 1, we will synthesize novel cocaine-based small
molecule haptens modified at the P3 site with various chemical linkers and linker lengths. We will then conjugate the
haptens to a self-assembling peptide domain using an orthogonal chemistry to produce peptide nanofibers that display the
haptens in a multivalent fashion. In aim 2, we will investigate antibody responses against modified cocaine haptens coupled
self-assembling peptides and the effect of linker chemistry and linker-length on the antibody titers. Mice will be immunized
with synthetic cocaine nanofiber vaccines and the production of anti-cocaine antibodies will be investigated using a primeboost regimen. Formulations that elicit high titers of anti-cocaine antibodies will be investigated for suppression of acute
cocaine-induced motor activity and cocaine-induced behavioral sensitization. Also, more broadly, completion of the
proposed work will integrate the fields of synthetic chemistry, nanotechnology, immunology, and addiction to significantly
impact human health.
4. Design, and Synthesis of a Peptide Hairpin Containing a Central Three-Residue Loop
The construction of a designed β-hairpin structure, containing a central three-residue loop has been successfully achieved in
the synthetic nonapeptide Boc-Leu-Phe-Val-DPro-LPro-DAla-Leu-Phe-Val-OMe (2). The design is based on expanding the
two-residue loop established in the peptide β-hairpin Boc-Leu-Phe-Val-DPro-LPro-Leu-Phe-Val-OMe (1). Characterization
of the registered β-hairpins in peptides 1 and 2 is based on the observation of key nuclear Overhauser effects (NOEs) in
CDCl3 and CD3OH. Solvent titration and temperature dependence of NH chemical shifts establish the identity of NH groups
involved in interstrand hydrogen bonding. In peptide 2, the antiparallel registry is maintained, with the formation of a DProL
Pro-DAla loop, stabilized by a 5→1 hydrogen bond between Val3 CO and Leu7 NH groups (C13, α-turn) and a 3→1
hydrogen bond between DPro4 CO and DAla6 NH groups (C7, γ-turn). NMR derived structures suggest that in peptide 2,
D
Ala6 adopts an αL conformation. In peptide 1, the DPro-LPro segment adopts a type II' β-turn. Replacement of DAla 6 in
peptide 2 by LAla in peptide 3 yields a β-hairpin conformation, with a central DPro-LPro two-residue loop. Strand slippage at
the C-terminus results in altered registry of the antiparallel strands.
5. Nanomaterials-based Vaccines for Cocaine Addiction, Mice Immunogenicity, and Behavioral Studies
The prevalence of cocaine abuse and dependence is a significant public health challenge in the USA with ~1.4 million
current users and accounts for nearly half of the illicit drug-related medical emergencies. Currently there are no FDA-
approved therapies for acute overdose or treatment of cocaine addiction. In recent years, vaccines that elicit effective anticocaine antibodies, which prevent cocaine penetration across the blood-brain barrier and interrupt its rewarding effects,
have been successful in animal models. However, a limiting factor is the degree and specificity of immunity evoked by the
addictive drug analog and small molecules like cocaine will not elicit an immune response unless administered with a
carrier protein or an adjuvant. The lack of effective clinically approved adjuvants has led to the development of selfadjuvanting vaccine platforms for various infectious and non-infectious diseases. Recently, self-assembling peptides that
assemble into β-sheet rich nanofibers have been shown to be effective for eliciting strong adaptive immune responses
against conjugated antigens without the need for any exogenous adjuvants. Here, we investigated the ability of selfassembling peptides to elicit antibody responses against small molecule drugs by linking cocaine analogs modified at the P3
site to the self-assembling peptide KFE8. Preliminary studies showed that vaccination with cocaine-bearing KFE8 peptide
nanofibers protected against cocaine-induced hyperactivity in mice.
6. Stereochemistry of Gabapentin and Several Derivatives: Solid State Conformations and Solution Equilibria
The role of γ-aminobutyric acid (GABA) as an inhibitory neurotransmitter has stimulated an enormous amount of activity
on the synthesis of GABA analogues as potential central nervous system agents. Gabapentin (1-(aminomethyl)
cyclohexaneacetic acid; Gpn) has been introduced as an anti-epileptic drug which is orally active. The mechanism of action
of Gpn remains a subject of considerable discussion. The branched chain amino acid metabolic pathways have been
implicated in several neurological disorders. Despite its structural simplicity, Gpn can exist in two distinct conformations
corresponding to the two interconvertible chair forms of the cyclohexane ring. Chemical structure illustrates the
consequences of the conformational interconversion for interaction at a potential receptor site. In principle, three major
elements of interaction can be considered; two involving electrostatic anchoring of the charged alkylamino and carboxylate
groups, while the third would be mediated by apolar (hydrophobic) contacts between the hydrocarbon ring and a potential
nonpolar group on the receptor. In this simplistic, three-point attachment model, originally formulated by Ogston, any chiral
receptor site would be readily able to distinguish between two possible chair conformations, suggesting that a specific
biological activity may be mediated by a single conformation. We have undertaken a systematic study of the structures of
Gpn and its derivatives in order to probe the factors which determine the conformational distributions in this class of 1,1disubstituted cyclohexanes. We discuss in this project, crystal structures of the free amino acid Gpn (1) and eight
derivatives, namely Ac-Gpn-NHMe (2). NMR evidence for the conformational interconversion and estimated free-energy
differences between the two forms in Gpn and its derivatives are also studied. The stereochemistry of Gpn and its
derivatives is also specially relevant in view of the growing interest in the use of γ-amino acid residues in the design.
COMPUTER SKILLS
• Operating Systems
: Windows, Macintosh, and Linux
• Applications
: MS Power Point, MS Excel, and MS Word
• Bioinformatics
: UniProt, PubMed, BLAST, and ABA
EXTRA CURRICULAR ACTIVITIES
• Volunteer in blood donation camp
• Science teacher in orphans school, and HIV/AIDS patients
• Counselor of psychotherapy and physiotherapy for HIV/AIDS patients
PERSONAL DETAILS
Gender
Citizenship
Marital Status
Male
Indian
Married
CONTACT ADDRESS
Dr. Rajagopal Appavu
515 1st Street, Apt #261
Galveston, TX77550
REFERENCES: Upon Request. Immunology, Virology, FACS, Elispot, HAI, Staining, Cytokines, Immunology, Virology, FACS, Elispot, HAI, Staining,
Cytokines