protein purification scientist

Zhishan (Tim) Huang, Ph.D.

Ossining, NY *****

Cell Phone: 914-***-****

E-mail: *******@*******.***

Experience

****-**** ********** ********* *************** Inc., Ridgefield, CT

Contract Scientist

• Expressed and purified numerous proteins (his-tagged proteins, GST-tagged proteins as well as proteins with no tag) using gel filtration, ion exchange, and affinity columns by AKTA chromatography ( Prime Plus, FPLC, Purifier and Explorer) for drug high throughput screening, structure research and antibody development in cardiovascular and immunological diseases.

• Purified monoclonal antibodies from hybridoma supernatants for therapeutic antibody development using AKTA, BioOptix and Filtration method.

• Conducted QC analysis (endotoxin measurement, dynamic light scattering, gel electrophoresis and staining, HPLC-SEC).

• Prepared samples for mass spectrometry analysis and biophysical analysis (AUC analysis).

• Antibody and peptide modifications

• Plasmid amplification and isolation in E. coli and protein synthesis using in vitro transcription and translation kit.

2006-2008 New York Medical College, Valhalla, NY

Postdoctoral Fellow and Lab Manager

• Investigated the mechanism how exogenous cytochrome c get into mitochondria in cytochrome c null cells using western blotting, immunostaining, fluorescence and electron microscopy, mitochondria oxygen consumption, COX activity, apoptosis assay (TUNEL) and cell proliferation (BrdU incorporation).

• Investigated whether caffeine can increase the function of septic hearts by measuring left ventricular pressure, perfusion pressure, heart rate and blood-gas in Langendorff system.

• Attempted to develop new drugs targeting ischemia-reperfusion injury in rat hearts using Langendorff system and immunohistochemical analysis.

• Animal handling techniques: animal anaesthetization, surgery, suture, post-surgery care and euthanasia.

• Developed new techniques and wrote new experimental protocols.

• Trained students and clinic fellows in various research projects.

2005-2005 National Institute of Child Health & Human Development, NIH, Bethesda, MD

Visiting Fellow (Apr. 2005-Aug. 2005)

• Expression of recombinant neurogranin in E. coli (plasmid extraction, transformation, induction, clone selection).

• Purification of recombinant neurogranin from E. coli crude lysates (acid precipitation ultracentrifugation, Amicon Pressure Filtration System, reverse phase HPLC with UV detector, lyophilization, purity analysis).

• Comparison of reduction activity of neurogranin and thioredoxin (insulin reduction assay).

• Investigation of the mechanism neurogranin reduction by NADPH in mice brain extracts and purification of potential upstream enzymes responsible for its reduction (heat precipitation, ultracentrifugation, Amicon Pressure Filtration System, FPLC with UV detector, non-reducing gradient gel and immunoblotting).

2000-2004 American Health Foundation, Valhalla, NY

Ph.D. Candidate, Graduate Research Associate

• Measured blood cotinine levels using ELISA in tobacco smoking study.

• Established an oral cancer model by treating Fisher rats with 4-NQO in drinking water.

• Measured thiols (glutathione, cysteine etc) in various rat tissues including tongue, lung, liver, brain and blood using both enzymatic and HPLC methods.

• Studied the inhibitory effect of caspase-3 glutathiolation in a cell free system.

• Detected glutathiolation of caspase-3 in vitro by biotin-labeling method and mass spectrometry.

• Identified the glutathiolation sites in recombinant caspase-3 by trypsin digestion and mass spectrometry.

• Detected glutathiolated caspase-3 in oral cancer as well as colon cancer by isoelectric focusing.

• Animal protocol and manuscript writing.

1998-1999 New York Medical College, Valhalla, NY

Rotating Ph.D. student

• Measured Ag85 titer in serum derived from TB patients using dot blot and ELISA.

• Studied the function of connexin 43 in astrocyte apoptosis.

• Measured PDE-3 and -4 mRNA levels using RNAase protection assay.

1997-1998 Nankai University, Tianjin, China

Research Assistant

• Investigated effects of organotin compounds on immune system in mice using blood cell count.

• Handling of research animals: anaesthetization, collection of animal blood, making blood smear slides and euthanasia.

• Housekeeping for laboratory animals.

Research Skills Molecular Biology: plasmid extraction and manipulation, bacteria transformation, agarose and PAGE gels, Northern blotting, RNAase protection assay, gel shift assay, densitometry.

Cell Biology: cell culture, cell transfection, light and fluorescence microscopy, sample preparation for electron microscopy, subcellular fractionation and differential centrifugation, apoptosis and cell proliferation assays, immunohistochemistry, sample preparation and data analysis for flow cytometry.

Immunology: Immunoassays (ELISA, RIA), Western blotting, dot blotting, immunoprecipitation immunohistochemistry.

Biochemistry: enzyme activity assays, protein expression and purification, HPLC, AKTA chromatography, BioOptix, SDS-PAGE, IEF, native gel, 2D gel, gel staining techniques, sample preparation and data analysis for mass spectrometry, dynamic light scattering, lyophilization.

Chemistry: IR, 1H NMR spectrum, melting and boiling points, reflective index, extraction, crystallization, distillation, thin-layer chromatography.

Physiology: animal surgery, isolated heart system (Langendorff preparation), AD Instrument Powerlab, blood gas analysis, mitochondria oxygen consumption.

Computer Skills Software: Microsoft Office, Adobe Reader and Photoshop, Reference Manager, Endnote, Sigma Plot, AD Instrument Chart 5, Image J, AlphaImager, NCSS statistics software, UNICORN 5.1, Vector NTI, Softmax Pro and EndoScan-V.

Sequence analysis (BLAST, Entrez ) and internet search (Pubmed and Medline)

Operation Systems: Windows (NT, XP, Vista and Server 2003)

Computer Language: C language

Education New York Medical College, Valhalla, NY 10595

Ph.D. in Experimental Pathology, May 2005

Nankai University, Tianjin, P. R. China

B.S. in Environmental Science, June 1997

Memberships Member of American Association for Cancer Research (2004-present)

Member of New York Science Academy (2003-present)

Research Interest Protein science

Cancer biology and apoptosis

Immunology and infectious diseases

Cardiovascular research

Redox biology

Publications

1. Verma R, Huang Z, Deutschman CS, Levy RJ. (2009) Caffeine restores myocardial cytochrome oxidase activity and improves cardiac function during sepsis. Crit Care Med. 37(4):1397-402.

2. Huang Z, Pinto JT, Deng H, Richie JP, Jr. (2008) Inhibition of caspase-3 activity and activation by protein glutathiolation. Biochemical Pharmacology. 75(11):2234-44.

3. Csiszar A, Labinsky N, Zhao X, Hu F, Serpillon S, Huang Z, Ballabh P, Levy RJ, Hintze TH., Wolin MS, Austad SN, Podlutsky A, Ungvari Z. (2007) Vascular O2.- and H2O2 production and oxidative stress resistance in two closely related rodent species with disparate longevity. Aging Cell. 6(6):783-97.

4. Huang Z, Komninou D, Kleinman, W, Pinto, JT, Gilhooly E, Calcognotto A and Richie JP, Jr. (2007) Enhanced levels of glutathione and protein glutathiolation in rat tongue epithelium during 4-NQO-induced carcinogenesis. International Journal of Cancer. 120(7):1396-401.

5. Landowski, CP, Godfrey HP, Bentley-Hibbert SI, Liu X, Huang Z, Sepulveda R, Huygen K, Gennaro ML, Moy FH, Lesley SA, Haak-Frendscho M. (2001) Combinatorial use of antibodies to secreted mycobacterial proteins in a host immune system-independent test for tuberculosis. J. Clin. Microbiol. 39:2418-24.

Manuscripts

1. Huang Z, Pinto JT, Deng H, Dai W, and Richie JP, Jr. Induction of caspase-3 glutathiolation during 4-nitroquinoline-N-oxide-induced tongue carcinogenesis in the rat. (Manuscript in preparation)

2. Khan AR, Seymour TL, Khan IR, Huang Z, Zhang S, Cohen MS, Spray TL, Deutschman CS, Gaynor JW, Levy RJ. Myocardial cytochrome c oxidase is inhibited after cardioplegic arrest. (Manuscript submitted)

Abstracts

1 Levy RJ, Huang Z, Edwards JG, Gao Q, Wolin MS. (2008) Functional import of exogenous mature cytochrome c into mitochondria of deficient cells. Experimental Biology Annual Meeting, FASEB J. 2008 22:1017.6. ( San Diego, CA, April 5-9, 2008)

2 Huang Z, Komninou D, Pinto JT, Richie JP, Jr. (2004) Regulation of caspase activity by glutathiolation. AACR Annual Meeting Proceeding 45: #5181. (Orlando, FL, March 27-31, 2004)

3 Richie JR Jr., Komninou D, Huang Z, Kleinman WA, Calcagnotto A, Schwartz JL. (2003) Enhanced protein glutathiolation in oral carcinogenesis. AACR Annual Meeting Proceeding 44: #6280. (Washington D.C., July 11-14, 2003)

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