a toms-turkiye He Heli licobacter cobacter pyl ylori ori %50 - PowerPoint PPT Presentation

a toms-turkiye

He Heli licobacter cobacter pyl ylori ori %50 %50

He Heli licobacter cobacter pyl ylori ori Peptic ptic Ul Ulcer ers St Stomac mach Cancer ncer

PEPTIC ULCERS 300,000 deaths

STOMACH CANCER 3rd leading cause

H. pylori

Penetr netrates tes under er muc ucous ous la layer er Hard d to to cur ure due ue to to ant ntibio ibiotic tic resista sistanc nce.

AIM AIM CURE CURE CA CANCER NCER

2. 2. ACI CID D 3. INCRE REAS ASED REP EPEL ELLENC ENCY MOTILI LITY TY 1. 1. ACI CID RES ESIST STANC ANCE 4. 4. SE SENSI SING H. pylori H. lori 5. KI 5. KILLING LING ERADIC ER ADICATIO TION N . .

BBa_K1639002 1. ACI 1. CID RES ESIST STANC ANCE H. H. pylori lori ER ERADIC ADICATIO TION N . .

Na Natur turally ally ac acid id res esista istant nt till ill pH pH 5

EXPECTED OBSERVED

Bacterial density in OD 600nm . Cell Viability 1200 1000 800 600 400 200 0 Pet45-b gadE Pet45-b gadE Pet45-b gadE Pet45-b gadE Pet45-b gadE pH 7.2 pH 5.0 pH 3.5 pH 2.5 pH 2.0 Beginning 3 min 1. hour 2. hour 5.hour 9.hour 13.hour

pH 7.2 pH 5.0 1500 1000 1000 500 500 0 0 (-) Control Pet45-b gadE Pet45-b gadE (-) Control Beginning 3 min 1. hour 2. hour Beginning 3 min 1. hour 2. hour 5.hour 9.hour 13.hour 5.hour 9.hour 13.hour

pH 2.5 pH 3.5 400 400 300 300 200 200 100 100 0 0 Pet45-b gadE (-) Control Pet45-b gadE (-) Control Beginning 3 min 1. hour 2. hour Beginning 3 min 1. hour 2. hour 5.hour 9.hour 13.hour 5.hour 9.hour 13.hour

pH 2.0 300 250 200 150 100 50 0 Pet45-b gadE Beginning 3 min 1. hour 2. hour 5.hour 9.hour 13.hour (-) Control

2. ACI 2. CID D REP EPEL ELLE LENCY NCY 3. INCRE REAS ASED MOTILI LITY TY 1. 1. ACI CID RES ESIST STANC ANCE 4. 4. SE SENSI SING H. pylori H. lori 5. 5. KI KILLING LING ER ERADIC ADICATIO TION N . .

BBa_K1639003 2. ACI 2. CID D REP EPEL ELLE LENCY NCY H. pylori H. lori ER ERADIC ADICATIO TION N . .

EXPECTED OBSERVED

2. ACI 2. CID D REP EPEL ELLE LENCY NCY 3. INCRE REAS ASED MOTILI LITY TY 1. 1. ACI CID RES ESIST STANC ANCE 4. 4. SE SENSI SING H. pylori H. lori 5. 5. KI KILLING LING ER ERADIC ADICATIO TION N . .

BBa_K1639004 BBa_K1639005 3. INCRE REAS ASED BBa_K1639006 MOTILI LITY TY H. H. pylori lori ERADIC ER ADICATIO TION N . .

a) A MUTANT FORM OF HNS

EXPECTED OBSERVED

b) Changing MotA& MotB to PomA&PotB

MotB PomB PotB (E. coli) (chimeric) (V. cholerae)

(-) Control HNS HNS-T108I PotB59 / PomA 0.25 24h 24h 24h 24h 0.40

Compar ared ed moti tili lities ties: : HNS-T108I > PotB59/PomA > > Wild type HNS > (-) control 10 Motility by fold 10 9 9 8 8 7 6 5 4 3 3 2 1 motility 0 HNS-T108I PotB59/PomA Wild type HNS (-) control

2. ACI 2. CID D REP EPEL ELLE LENCY NCY 3. INCRE REAS ASED MOTILI LITY TY 1. 1. ACI CID RES ESIST STANC ANCE 4. 4. SE SENSI SING H. pylori H. lori 5. 5. KI KILLING LING ER ERADIC ADICATIO TION N . .

BBa_K1639014 BBa_K1639000 BBa_K1639001 4. 4. SE SENSI SING H. H. pylori lori ER ERADIC ADICATIO TION N . .

BBa_K1639014 BBa_K1639000 BBa_K1639001 4. 4. SE SENSI SING H. H. pylori lori ER ERADIC ADICATIO TION N . .

And Logic Gates

Toehold-RNA Switches

The presence of Trigger RNA in the cell unwinds the hairpine structure. Then RBS sets free and causes the production of TEV Protease

We used GFP instead of TEV protease

IPTG (-) IPTG (+) pColA ToeHold GFP + Trigger RNA pColA ToeHold GFP

pColA-ToeHold GFP pColA-ToeHold GFP + Trigger RNA IPTG - - + +

Cell Lysate Fold Change Avg 300 250 200 150 100 50 0 − + − + Toehold Toehold+Trigger

2. ACI 2. CID D REP EPEL ELLE LENCY NCY 3. INCRE REAS ASED MOTILI LITY TY 1. ACI 1. CID RES ESIST STANC ANCE 4. SE 4. SENSI SING H. pylori H. lori 5. 5. KI KILLING LING ER ERADIC ADICATIO TION N . .

BBa_K1639007 BBa_K1639008 H. H. pylori lori 5. 5. KI KILLING LING ER ERADIC ADICATIO TION N . .

Pexig xiganan anan A nt nti i M icr icrobial obial P ept ptide ide

EXPECTED OBSERVED

EXPECTED OBSERVED

‘ If you achieved just one of your goals that would be exciting enough. ’ Barry ry J. Ma . Marsh shall all 2005 Nobel Prize laureate in Physiology or Medicine

STOMA OMACH CH CA CANCER NCER BBa_K1639015 BBa_K1639016 TR TREA EATMENT TMENT BBa_K1639010 BBa_K1639011 BBa_K1639009 6. CA 6. CANCE CER SWITCH

miRNA switch working mechanism

Human Practices

Going public: iGEM for little kids

Outreach: Street Interview Interviewed with; 1)Patients 2)Doctors 3)People

iGEM Meetup: Synbio Day

A very new approach to medicine; VIRTUAL HOSPITAL where synthetic biology meets medicine in future

Collaborations Mentored AUC Turkey iGEM team Prepared a VH video of UCL iGEM Team’s project ‘ Mind the Gut’ Connected our team to the iGEM Rhizi (created by Paris Bettencourt iGEM) Held a meetup with 3 other regional teams Joined to the iGEM Academy YouTube channel (Created by Tinity iGEM)

Safety: Suicide Switch BBa_K1639022

ACHIEVEM HIEVEMENTS ENTS

Our ur No Novel el Pl Plas asmids mids pSB SB1C3 1C3-T7 T7 (ATOMS OMS) pColA lA (ATOMS OMS)

MAIN REFERENCES [1] Ma, Z., Gong, S., Richard, H., Tucker, D., Conway, T., & Foster, J. (n.d .). GadE (YhiE) activates glutamate decarboxylase-dependent acid resistance in Escherichia coli K-12 . Molecular Microbiology, 1309-1320. [2] Croxen, M., Sisson, G., Melano, R., & Hoffman, P. (2006). The Helicobacter pylori Chemotaxis Receptor TlpB (HP0103) Is Required for pH Taxis and for Colonization of the Gastric Mucosa . Journal of Bacteriology, 2656-2665. [3] Nishino, Y., Onoue, Y., Kojima, S., & Homma, M. (2015). Functional chimeras of flagellar stator proteins between E. coli MotB and Vibrio PomB at the periplasmic region in Vibrio or E. coli. MicrobiologyOpen , 323-331. [4] Donato, G., & Kawula, T. (1998). Enhanced Binding of Altered H-NS Protein to Flagellar Rotor Protein FliG Causes Increased Flagellar Rotational Speed and Hypermotility in Escherichia coli . Journal of Biological Chemistry, 24030-24036. [5] Green, A., Silver, P., Collins, J., & Yin, P. (n.d.). Toehold Switches: De-Novo-Designed Regulators of Gene Expression . Cell, 925-939. [6] Lewis V Wray Jr., Jill M Zalieckas, Susan H Fisher. Bacillus subtilis Glutamine Synthetase Controls Gene Expression through a Protein- Protein Interaction with Transcription Factor TnrA . Cell Volume 107, Issue 4, 16 November 2001, Pages 427 – 435 [7] Vendeville, A., Winzer, K., Heurlier, K., Tang, C., & Hardie, K. (n.d.). Making 'sense' of metabolism: Autoinducer-2, LUXS and pathogenic bacteria . Nature Reviews Microbiology Nat Rev Micro, 383-396. [8] Zhang XL, Jiang AM, Ma ZY, Li XB, Xiong YY, Dou JF, Wang JF. The synthetic antimicrobial peptide pexiganan and its nanoparticles (PNPs) exhibit the anti-helicobacter pylori activity in vitro and in vivo. Molecules 2015, 20(3), 3972-3985; doi:10.3390/molecules20033972 [9] Zhao, C., Dwyer, M., Yu, A., Wu, Y., Fang, S., & Middelberg, A. (2015). A simple and low-cost platform technology for producing pexiganan antimicrobial peptide in E. coli . Biotechnol. Bioeng. Biotechnology and Bioengineering, 957-964.

INSTR IN STRUCT UCTORS ORS & A & ADVISORS VISORS Prof. Dr. Esra Gündüz Prof. Dr. Mehmet Gündüz Assist. Prof. Muradiye Acar Assist . Prof. Dr. Sultan Çiftçi Yılmaz Assist. Prof. Dr. Ömer Faruk Hatipo ğ lu Dr. Sadık Çi ğ dem Ay ş e Çelik Burak Yılmaz Mustafa Semih Elitok

ACK CKNO NOWLEDGEMENTS WLEDGEMENTS Dr. Akihiko Ishijima We’d like thank you to Dr.Akihiko for providing us amino acid sequence of PotB59 protein Dr. Andrea Schuller For their big support and advices about DAMP4 and Pexiganan’s mechanism Dr. Chunxia Zhao We’d also thank you to Dr.Chunxia for answering our questions about Pexiganan Dr. Bernard Binetruy Special thanks to Dr.Bernard for providing of CAGop promoter’s sequence

SP SPONSORS ONSORS

Thanks for your patience

EXTRA SLIDES

TlpB GadE

Potb59/PomA HNS-T108I

pAlst-TnrA pLsr-LsrR

Toehold Switch Trigger RNA

DAMP-Pexiganan TEV Protease