ENGINEERING AN EDIBLE PROBIOTIC CONSORTIUM TO REGULATE APPETITE

AND DIGESTION

COLUMBIA NYC

Jacky Cheung, Samuel Magaziner, Suppawat Kongthong, Hudson Lee, Kenya Velez

THE RISE OF METABOLIC DISEASES

CHANGES IN CALORIC INTAKE PER CAPITA PER

DAY OVER TIME IN THE U.S

COMPARISON OF AVERAGE CALORIC INTAKE PER

CAPITA AROUND THE WORLD

OBESITY & DIABETES MEDICAL EXPENSES QUALITY OF LIFE

FOOD AS THE SOLUTION – NOT THE PROBLEM

HISTORICAL AND PROJECT SALES OF YOGURT IN THE UNITED STATES WHAT ARE PROBIOTICS?

Live Bacteria & Yeasts

That Improve Health

DIGESTIVE FUNCTIONS

IMMUNE SYSTEM

FUNCTIONS

SUPPORT FRIENDLY

BACTERIA

“Let food be thy medicine and medicine be they food” - Hippocrates

GOAL: ENGINEER A PROBIOTIC FOOD THAT COUNTERS OBESITY AND DIABETES

METHOD TO COUNTER OBESITY AND DIABETES

GLUCAGON-LIKE PEPTIDE I (GLP-I)

INSULIN LEVELS

SATIETY LEVELS

PEPTIDE YY (PYY)

APPETITE LEVELS

GI MOVEMENT

GHRELIN

APPETITE LEVELS

CARDIOVASCULAR HEALTH

ENGINEERING AN EDIBLE PROBIOTIC TO SECRETE GUT PEPTIDES TO REGULATE APPETITE AND DIGESTION

E. C

oli N

issle

Lact

obaci

llus

Reu

teri

phoA

pelB

Lp_3050

M6

GLP-1

Ghrelin PYY

Signal Peptides

Sig

nal Pe

ptid

es

(SPs)

x

Gut Pep

tid

es

(GPs)

EXPERIMENTAL SETUP: SIGNAL PEPTIDE SECRETION

Anhydro-Tetracycline (or

Nisin in Lactobacillus model)

pTet HormoneSP His-TagLysate

1X101

1X10-1

1X10-2

1X102

1X101

1X10-1

Supernatant

His-tagged

Protein

Concentrate

supernatant +

lyse cells

Grow-up, spin

down cell pellet,

extract

supernatant

Blot

RESULTS: HORMONES (NO SIGNAL PEPTIDE)

Ghrelin PYYGlp-1

Note: We expect to see no secretion

Lysate (Left) 1x101 Supernatant (Right) 1x102

Lysate (Left) 1x10-1 Supernatant (Right) 1x101

Lysate (Left) 1x10-2 Supernatant (Right) 1x10-1

RESULTS: SUCCESS WITH PHOA-GLP1 (IMPORTANCE OF INDUCTION TIME)

Lysate (Left) 1x101 Supernatant (Right) 1x102

Lysate (Left) 1x10-1 Supernatant (Right) 1x101

Lysate (Left) 1x10-2 Supernatant (Right) 1x10-1

PhoA-Glp1(36hr induction)PhoA-Glp1 (12hr Induction)

Lysate 1x101

Supernatant 1x101

Supernatant 1x10-1

Note: Same construct, Wildly different Results; also

improvement of previous biobrick BBa_K817000

RESULTS: SUCCESS WITH PELB

Lysate 1x101

Supernatant 1x101

Supernatant 1x102

PelB-Glp1 (36hr) PelB-PYY (36hr)

Here we demonstrate the generalizable

nature of signal peptide secretion

IMPLEMENTING A CONSORTIUM TO PROVIDE REGULATIONS AND SAFETY MECHANISM FEATURES

COMMUNICATION

REGULATION

SAFETY

OUR SYSTEM IS TO BE

TIGHTLY REGULATED AS A

SAFETY PRECAUTION

THE SYSTEM: SAFETY, REGULATION, COMMUNICATION, AND SECRETION MEET

THE SYSTEM BREAKDOWN: THE AHL FACTORY

THE SYSTEM BREAKDOWN: TARGET SECRETION

THE SYSTEM BREAKDOWN: TIME-DELAYED CELL LYSIS (BBA_K1848006)

RESULTS

• Timer-Lysis Cassette system submitted as

biobrick (BBa_K1848006)

• System successfully cloned in

• Time constraints prevented its testing

• However…

1 32

Heat whole milk to

185ºF

Cool to 115ºF and

add engineered L.

reuteri + yogurt

starter culture

Incubate at 37ºC for

8 hr and refrigerate

IN PRACTICUM: YOGURT

Note: Yogurt was not intended for human consumption, nor did it leave the lab

SURVEYING THE PUBLIC• Online survey

• Polled public on GMOs and probiotics, to inform design of safety measures and yogurt

products

• Informed survey takers about probiotics and GMOs

• Twitter and blog

Comfort level in consuming probiotics (On scale of 1-5) before

informing the survey takers about probiotics

Comfort level in consuming probiotics (On scale of 1-5)

after informing the survey takers about probiotics

THE FUTURE: YOGHURT (OR LOOKING WHEY INTO THE FUTURE)

• Going Forward:

1. Finish screen of submitted quorum sensing system

2. Move system into L. reuteri

3. Test active system’s ability to function within yogurt

culture

4. Screen more hormones

• The Foreseeable future:

• Catalog of peptide secreting yogurt

• Biotech branch based around personalized consumable

medicines

IN SUMMATION…

• We managed to:

1. Submit 6 bio-bricks (5 of which are well-

characterized) (BBa_K1848001-006)

2. Successfully secrete 3 gut hormones and 2 signal

peptides in E. coli

3. Develop a promising system quorum sensing based

secretion and lysis

4. Move an engineered L. reuteri system in yogurt

5. Poll on public opinion of probiotics and willingness to

consume GMOs

6. Have an amazing summer and lay the foundation for

future iGEM teams from our University

Student Team MembersThe Whole Team

ATTRIBUTIONS AND ACKNOWLEDGMENT

• Undergraduate Team Members: Suppawat Kongthong, Jacky Cheung, Kenya

Velez, Hudson Lee, Samuel Magaziner

• Mentors: Nathan Johns, Sway Chen, and Sonja Billerbeck

• PIs: Harris Wang, Virginia Cornish, Ken Shepard, Dana Pe’er

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