Team:Hong Kong UCCKE/Description

Where did our idea come from?

1% of the World’s population are suffering from gout and it is becoming more prevalent, solely in the U.S, 8.3 million people[1] were affected. Our team’s PI’s family have been suffering from the disease for generations as well. As his students, as well as a group of passionate high school science students, we were inspired to find new detection methods and even a cure for gout.

What is Gout?

Gout is a form of arthritis, caused by the elevated levels of uric acid in the blood. At high levels, uric acid crystallizes and accumulates in synovial fluid, leading to massive pain, and a red, tender, hot, swollen joint. [2]
The food we intake everyday such as steak, organ meats and seafood contains purine. The purines we intook will be catalyzed by several enzymes into uric acid (as shown on the pathway below). However, we, as homo sapiens, lack uricase, as known as urate oxidase, which catalyzes uric acid into allantoin.

Pathway from purine to Uric acid Pathway from purine to Uric acid

Normally, uric acid dissolves in our blood and passes through our kidneys into our urine. But sometimes the body produces too much uric acid, or when the kidneys are unable to excrete sufficient amounts of uric acid efficiently. Along with the low solubility of uric acid in water (0.6mg/100ml), this can result in a build up of tiny needle-sharp crystals in and around joints.

Uric acid accumulate in toe Uric acid accumulate in toe
Feet of patient suffering from gout Feet of patient suffering from gout
Uric acid excreted by kidney Uric acid excreted by kidney

What is our plan?

There are three main sub-projects. We wish to detect patients’ gout symptoms, and to cure it by preventing excess uric acid from accumulating in the body. Several parts have been designed to achieve the above goals.

1.Blood uric acid level detector

Firstly, we hope to make a uric acid detector, which is Part BBa_K2197300. It encodes HucR, a repressor, and a GFP gene. [3]Working with KRAB, a repressor enhancer, and hucO, an operative site for the above repressor, the first session of our part acts as a uric acid concentration sensitive promoter. The promoter controls the expression of GFP. Engineered e.coli encodes part BBa_K2197300. By adding blood samples to the culture medium of e.coli, it expresses different levels of GFP. By measuring the GFP level with a plate reader, uric acid concentration of the sample can be estimated. This part ensures a rapid detection of the uric acid concentration inside the patient's body.

Expression of GFP in presence of uric acidExpression of GFP in presence of uric acid

2.Uric acid decomposition

We want to make a uric acid decomposer, Part BBa_K2197400. Having the same promoter as the previous one, this cell produces urate oxidase, an enzyme catalysing the oxidation of uric acid. First to 5-hydroxyisourate, then to allantoin
(Uric acid + O2 + H2O → 5-hydroxyisourate + H2O2
5-hydroxyisourate + H2O → allantoin + CO2)
But the human gene for urate oxidase is nonfunctional. In the future, it is hoped a capsule enclosing this engineered cell can be made so that it can be taken in by patients and produce urate oxidase when it senses uric acid.

UOX in human bodyUOX in human body

3. Uric acid transporter

We made a uric acid transporter, YgfU[4], which is an imitation of Glut9. Glut9, as known as olute-carrier 2A9, plays a major role in urate homeostasis by its dual role in urate handling in the kidney and uptake in the liver. For YgfU, it is an E.coli-originated uric acid transporter. Having the same promoter as the two previous projects, in the presence of uric acid, YgfU is expressed, thus, the cell can absorb uric acid.

The protein Glut9a, Glut9b in kidneyThe protein Glut9a, Glut9b in kidney

Reference

[1]: Gout Becoming More Common in U.S. - WebMD

[2]: Martin Underwood. Diagnosis and management of gout. BMJ. 2006 Jun 3; 332(7553): 1315–1319.

[3]: Christian Kemmer, Marc Gitzinger, Marie Daoud-El Baba, Valentin Djonov, Jorg Stelling & Martin Fussenegger (2010) Self-sufficient control of urate homeostasis in mice by a synthetic circuit. Nature Biotechnology doi: 10.1038/nbt.1617

[4]: Martin Underwood. Diagnosis and management of gout. BMJ. 2006 Jun 3; 332(7553): 1315–1319.

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