About OMICS Group OMICS Group International is an amalgamation of Open Access publications and worldwide international science conferences and events. Established in the year 2007 with the sole aim of making the information on Sciences and technology ‘Open Access’, OMICS Group publishes 400 online open access scholarly journals in all aspects of Science, Engineering, Management and Technology journals. OMICS Group has been instrumental in taking the knowledge on Science & technology to the doorsteps of ordinary men and women. Research Scholars, Students, Libraries, Educational Institutions, Research centers and the industry are main stakeholders that benefitted greatly from this knowledge dissemination. OMICS Group also organizes 300 International conferences annually across the globe, where knowledge transfer takes place through debates, round table discussions, poster presentations, workshops, symposia and exhibitions. About OMICS Group Conferences OMICS Group International is a pioneer and leading science event organizer, which publishes around 400 open access journals and conducts over 300 Medical, Clinical, Engineering, Life Sciences, Pharma scientific conferences all over the globe annually with the support of more than 1000 scientific associations and 30,000 editorial board members and 3.5 million followers to its credit. OMICS Group has organized 500 conferences, workshops and national symposiums across the major cities including San Francisco, Las Vegas, San Antonio, Omaha, Orlando, Raleigh, Santa Clara, Chicago, Philadelphia, Baltimore, United Kingdom, Valencia, Dubai, Beijing, Hyderabad, Bengaluru and Mumbai. YEDITEPE UNIVERSITY GENETICS AND BIOENGINEERING DEPT. Assist. Prof. Dr. Hüseyin Çimen [email protected] YEDITEPE PROTEOMICS and MASS SPECTROMETRY LABORATORY ISTANBUL, TURKIYE OMICS Aug 4th,2014 About me… Undergraduate (BSc): 2006 Molecular Biology and Genetics Middle East Technical University, TURKEY Graduate (PhD): 2012 Biochemistry, Microbiology and Molecular Biology (BMMB) Eberly College of Science, Pennsylvania State University, PA Research: 2011 - 2012 Biochemistry John C. Edwards Medicine School Marshall University, WV YEDITEPE UNIVERSITY STEM CELL AND GENE THERAPY EXCELLENCE CENTER Molecular Biology ve Genetics Lab Microbial Genetics Lab Plant Biotechnology Lab Bioinformatics Lab Biomaterials Lab Mol. Diagnostic Lab Functional Genetics Lab Protemics Lab Nanobiotechnology Lab Imaging Lab YEDITEPE PROTEOMICS and MASS SPECTROMETRY LABORATORY Discovery-based Targeted Proteomic Studies: Gel-based and/or off-gel protein sample preparation and purification nano-HPLC Allison Doerr, Nature Methods 10 (2013) Thermo Dionex UltiMate 3000 nanoESI-qTOF Bruker Compact •Protein ID •Protein structure, function, folding and interaction analysis •Post-translational modification (PTM; acetylation) analysis of complex biological samples •Biomarker discovery and characterization Biochemistry Proteomics Stanley Fields, Science 291 (2001) Targeted Proteomics Katie Vicari,, Nature Methods 10 (2012) Reproducible - Quantitative Highly Sensitive Protein ID and Biomarker Discovery Where is Waldo? Vancouver Canucks Fan Zone, 2011 http://www.gigapixel.com/image/gigapan-canucks-g7.html Where is Waldo? Where is Waldo? Where is Waldo? HERE? Vancouver Canucks Fan Zone, 2011 http://www.gigapixel.com/image/gigapan-canucks-g7.html Mitochondrial functions and cancer: Elevated glycolytic metabolism and oxygen use in cancer cells Douglas C. Wallace, Nature Reviews Cancer Electron Transport Chain COMPLEX nDNA mtDNA I 39 7 II 4 0 III 10 1 IV 10 3 V 16 2 H++ H++ H+ HH H+ ND2 Inner Membrane ND1 Cyt C e- e- ND3 ND4 ND6 ND4L ND5 e Q e- e- e- eMatrix ATP6 Cyt b ATP8 COX I COX II COX III +H+ H2OH H+H+- +`+` HH H+NAD+ Cyt C e- e- - +H + 2O O2 OH H H++H H O2 ADP O2 ATP .- .O2 O2 NADH Krebs Cycle Peter S. Rabinwitch after Mandavilli et al, Mutation Research 509 (2002) 127–151 Regulation of Oxidative Phosphorylation Comlexes Cyt C ND2 Inner memb. ND1 ATP6 ND3 Cyt b ND4 ND6 ND4L ND5 ATP8 COX I COX II COX III Q ac-NDUFA9 H2O Matrix ac-SdhA NAD+ NADH +H+ ADP Krebs Cycle Pi ATP Mitochondrial Proteome: >20 % acetylation (Kim SC et.al 2006) Lysine-NH2 Acetyl-CoA Acetyl-CoA CoA Lysine-NH-CO-CH3 1 Protein Protein 2 NAM O-acyl-ADP-ribose 1 2 NAD+ Acetylated GCN5L1 SIRT3, SIRT4, and SIRT5 Cimen H et al. (2010), Ahn BH et al. (2008), Scott I et al. (2012), and Kim SC et al. (2006). REFERENCE CENTRE of BORON STUDIES • BOREN (National Boron Research Institute, TURKIYE) – Efficiency and spectrum of scientific studies on boron • APPLICATION: – Glass, ceramic, flame retardants, cement, metallurgy, energy, cleaning, bleaching, agriculture, cosmetics, health – BNCT: boron neutron capture therapy in cancer REFERENCE CENTRE of BORON STUDIES • Concentrated Borates – Ulexite – Tincal – Colemanite • Refined Borates – – – – – – – – Borik Acid Boraks Pentahidrat Boraks Dekahidrat Etibor- 48 Sodyum Perborat Monohidrat Sodyum Perborat Tetrahidrat Borax Anhydroust Boron Oxide • Speciality Boron Products – Tarımbor (Agricultural Boron) – Ahsapbor (Wood Boron) – Zinc Borate – Cellulosic Insulation Material – Boron Nitride – Boron-Cement (Boron-AddedCement) – Elementel Boron – Sodium Borohydride – Trimetyl Borate – Boron Carbide BORON vs NAD+ Interaction between sodium borate and NAD+ may affect Sirtuins, NAD+ dependent deacetylases. (Hunt CD, 2012) BORON vs NAD+ Hep3B cells incubated with Sodium Borate supplemented media 72 h incubation with Serum starvation for 24 h Cell harvest and sample preparation Cellular, metabolic, and proteomic analyses NaB treatment decreased cell proliferation rate 1000 Con NaB 2 3 Cell Viability (%) 800 600 400 200 0 1 4 Time (Days) Percent cell viability measured through metabolic activity of cultured HEP3B cells treated with NaB. NaB treatment decreased overall protein acetylation A B NaB treatment (μg /ml) Relative Acetylation (%) 0 120 100 80 60 40 20 0 15 Acetyl-K 0 15 NaB treatment (μg /ml) GAPDH IB analysis demonstrated >10% decrease in whole cell acetylome NaB treatment decreased overall mitochondrial protein acetylation A B NaB treatment (μg /ml) Relative Acetylation (%) 0 120 100 80 60 40 20 0 15 Acetyl-K 0 15 NaB treatment (μg /ml) HSP60 IB analysis demonstrated >10% decrease in mitochondrial acetylome NaB treatment enhanced SIRT3 activity Relative SIRT3 Activity (%) 140 120 100 80 60 40 20 0 0 15 NaB treatment (μg /ml) NaB – NAD interaction elevated SIRT3 activity. CycLex® SIRT3 Deacetylase Fluorometric Assay Kit NaB treatment affects metabolism NAD+ / NADH Ratio Relative Level (%) 300 * 250 200 150 100 50 0 0 15 NaB treatment (μg /ml) NaB – NAD interaction elevates NAD+/NADH ratio. NaB treatment reduced OXPHOS complexes A B CV CIII CIV * Relative OXPHOS (%) * * CII 0 CI * NaB treatment (μg /ml) 15 CV * C III C IV C II 0 15 0 15 0 15 0 15 NaB treatment (μg /ml) 0 15 CI HSP60 NaB reduced mitochondrial biogenesis. OXPHOS Ab cocktail: ATPSA (CV), UQCRC2 (CIII), MTCO1 (CIV), SDHB (CII), NDUFB8 (CI) NaB treatment lowered ROS production Relative ROS Level (%) 140 120 100 80 60 40 * 20 0 0 15 NaB treatment (μg /ml) NaB reduced ROS generation (>70%). Abcam DCFDA Cellular ROS Detection Assay Kit Summary NaB Treatment (15 μg/ml) NAD+/NADH 103% ACETYLATION Cell Viability WCL SIRT3 activity Cancer cell proliferation hampered by BORON MITO BORON treatment enhances mitophagy in order to increase mitochondrial quality upon deacetylation. Future Prospects and Acknowledgement • Mitochondrial structure analysis with electron microscopy • Drug supplementation along with boron • Specific column development for boron-interacting proteins LAB MEMBERS: Graduate: MSc Berna USTUNER MSc Eray Esendir Special Thanks: Yeditepe SAHIN Lab members Firdevs Cansu ATILGAN Bihter YAVUZ Technician: Binnur Herand KIRATLI FUNDING: Yeditepe UNIVERSITY YEDITEPE GENETICS AND BIONEGINEERING PROTEOMICS and MASS SPECTROMETRY LAB. THANKS Department web: www.genetik.yeditepe.edu.tr OMICS Aug 4th,2014 Yeditepe Biyoteknology Society: yeditepebiotech.com fb.com/pages/Yeditepe-University-BiotechnologySociety Let Us Meet Again We welcome you all to our future conferences of OMICS Group International Please Visit: www.omicsgroup.com www.conferenceseries.com www.proteomicsconference.com
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