Background
Diabetes mellitus, hypertension and intestinal diseases are among the most common diseases worldwide, and their frequency increases with increasing age, seriously affecting the quality of life and health of human beings. By 2045, IDF projections show that 1 in 8 adults, approximately 783 million, will be living with diabetes, an increase of 46%. Among them, ore than 95% of people with diabetes have type 2 diabetes. More importantly, type 2 diabetes mellitus and hypertension overlap in the population.
The vast majority of patients with diabetes and hypertension must take drugs for a long time or even for life, and short-acting drugs should be supplemented in time.
Therefore, in order to make up for the shortcomings of traditional medicine, our team adopts Synbiotic Therapy, which has the advantages of high safety, good stability, strong adaptability, high targeting, long half-life and so on. We used Zymomonas mobilis as the carrier. It is a strain widely used in various industries. What’s more, it has a unique hopane membrane structure, strong tolerance to high sugar, alcohol and organic acids, and strong intestinal colonization ability. As a probiotic, it can be used to produce fructans and fructooligosaccharides. In addition, the genome of Fermentomonas motility has good stability and strong operability, so it can better help us achieve our goals.
technical route
1)cSAT- Product expression: The team members added a cleavable self-aggregable cSAT system to facilitate the correct expression of the protein, and then inserted the modified expression plasmid pTZ32a. The f1 ori of pET32 was replaced by zymo_replication. Expressed in Zymomonas mobilis.
2)Acidity sensor: Considering that polypeptides with hypoglycemic or antihypertensive effects are easily degraded during the fermentation process of Zymomonas mobilis or when entering gastric juices, we reconstructed a set of pH sensors. When the ambient pH is less than 5.5, the rcfB promoter senses the change in acidity, as a result, inhibiting downstream gene expression.
3)Antibiotic replacement: In order to ensure that our products are harmless to human health, the team members replaced the ampicillin resistance gene in the expression vector pTZ32a with the Nisin resistance gene nisl, and constructed a food-grade expression vector.
4)Tandem repeat active peptide: To enable the active peptide to be directly secreted in the human gut and hydrolyzed into the active peptide monomer enterokinase, the team members inserted the fusion protein gene sequence before the extracellular display of the protein, and tandem repeats of the 4×GLP-1 and 5×LV design were used to construct a polymer.
5)Bacterial coating: We use chitosan and sodium alginate as effective biofilms, and the coated bacteria are able to resist the acidic environment of the stomach, pass through various obstacles, and eventually reach the intestine.
6)Safe suicide switch design: After a period of time, probiotics are excreted with the stool. Based on the differences between the human body and the natural environment, we designed a suicide mechanism of glucose starvation. Due to the lack of glucose in the environment, resulting in bacterial autolysis and death.
expected result
When the bacteria involved in the research team enter the gut, if the environmental pH is greater than 5.5, GLP-1 is expressed and secreted, which binds to the GLP-1 receptor on β-islet cells through the blood to eventually produce insulin. Angiotensin-converting enzyme inhibitor (ACEi) LV recognizes and inhibits ACE enzyme activity in this environment, leading to vasodilation and decreased blood pressure. After a period of time, the probiotics will be excreted in the stool. Due to the lack of glucose in the environment, the bacteria autolysis and death.