Arginine supplementation to improve efficiency in gas fermenting acetogens
The invention provides methods for improving efficiency of fermentation by arginine supplementation, and genetically modified bacterium for use therefor. More particularly the invention provides methods for (i) increasing the production ATP intensive products with arginine supplementation, (ii) increasing utilization of arginine by a C1-fixing bacterium; and (iii) providing C1-fixing bacterium with optimized arginine de-aminase pathways.
1. A method for improving efficiency of arginine co-utilization with one or more gaseous substrates selected from the group consisting of CO, H 2 and CO 2 , the method comprising culturing a genetically engineered C1-fixing bacterium comprising one or more genetic modifications selected from the group consisting of:
i) a disruptive mutation in an arginine transporter;
ii) overexpression of one or more endogenous enzymes selected from the group consisting of arginine deiminase (EC 3.5.3.6), ornithine carbomyltransferase (putrescine carbomyltransferase) (EC 2.1.3.3), carbamate kinase (EC 2.7.2.2), ornithine racemase (EC 5.1.1.12), ornithine aminomutase (EC 5.4.3.5), and 2,4-diaminopentanoate dehydrogenase (EC 1.4.1.12);
iii) expression of one or more mutated endogenous enzymes selected from the group consisting of arginine deiminase (EC 3.5.3.6), ornithine carbomyltransferase (putrescine carbomyltransferase) (EC 2.1.3.3), carbamate kinase (EC 2.7.2.2), ornithine racemase (EC 5.1.1.12), ornithine aminomutase (EC 5.4.3.5), and 2,4-diaminopentanoate dehydrogenase (EC 1.4.1.12); and
iv) expression of one or more exogenous enzymes selected from the group consisting of arginine deiminase (EC 3.5.3.6), ornithine carbomyltransferase (putrescine carbomyltransferase) (EC 2.1.3.3), carbamate kinase (EC 2.7.2.2), ornithine racemase (EC 5.1.1.12), ornithine aminomutase (EC 5.4.3.5), and 2,4-diaminopentanoate dehydrogenase (EC 1.4.1.12) wherein arginine is a sole nitrogen source.
2. The method of claim 1 , further comprising one or more genetic modifications selected from the group consisting of (i) disruptive mutation of regulatory elements and (ii) replacement of operator binding sites or native promoters with constitutive or synthetic promoters.
3. The method of claim 2 , wherein the disruptive mutation is a knock-out of arginine repressor argR.
4. The method of claim 2 , wherein the replacement is a replacement of an arginine deaminase pathway operon promoter with a constitutive or synthetic promoter.
5. The method of claim 1 , wherein arginine is provided to the culture in excess of a cellular requirement of the culture.
6. The method of claim 1 , wherein the C1-fixing microorganism is a Clostridium bacterium.