Genetically modified microorganisms which are resistant to infection by bacteriophages and that retain their kinetic parameters and methods of making the same.

A cyanobacterium transformed using a gene transfer vector. The gene transfer vector includes: a first homologous recombination region homologous to a 5′ side of a target DNA of the cyanobacterium; a second homologous recombination region homologous to a 3′ side of the target DNA; and a DNA fragment introduced into a portion sandwiched between the first homologous recombination region and the second homologous recombination region. A total length of the first homologous recombination region and the second homologous recombination region is 5 kbp or more. The cyanobacterium has a DNA fragment transferred between the 5′ side of the target DNA and the 3′ side of the target DNA.

Disclosed are a ketoreductase mutant and a method for producing a chiral alcohol. The ketoreductase mutant has an amino acid sequence obtained by the mutation of the amino acid sequence shown in SEQ ID NO: 1, and the mutation includes a mutation siteK200H. In the present disclosure, the mutant obtained by mutation takes a ketone compound as a raw material, the chiral alcohol may be efficiently produced by stereoselective reduction, and the stability is greatly improved, which is suitable for popularization and application to the industrial production of the chiral alcohol.

The present invention provides engineered methionine gamma lyase polypeptides and compositions thereof. The engineered methionine gamma lyase polypeptides have been optimized to provide improved thermostability, protease stability, and stability under a range of pH conditions, including acidic (pH<7) conditions. The present invention also relates to the use of the compositions comprising the engineered methionine gamma lyase polypeptides for therapeutic purposes.

Microbial consortia exert great influence over the physiology of humans, animals, plants, and ecosystems. However, difficulty in controlling their composition and population dynamics have limited their application in medicine, agriculture, biotechnology, and the environment. The approach disclosed herein provides an effective method to dynamically control population compositions in microbial consortia, which we demonstrate in the context of co-culture fermentations for chemical production. Co-culture fermentations can improve chemical production from complex biosynthetic pathways over monocultures by distributing enzymes across multiple strains, thereby reducing metabolic burden, overcoming endogenous regulatory mechanisms, or exploiting natural traits of different microbial species. However, stabilizing and optimizing microbial sub-populations for maximal chemical production remains a major obstacle in the field. An optogenetic circuit, called OptoTA, is disclosed for regulating a toxin-antitoxin system, which enables tunability of, e.g., Escherichia coli growth using only blue light. With the disclosed system, one can control population ratios of co-cultures of, e.g., E. coli and Saccharomyces cerevisiae containing different metabolic modules of biosynthetic pathways. Results reveal that intermediate light duty cycles improve chemical production by establishing optimal co-culture populations.

The disclosure is in the technical field of synthetic biology and metabolic engineering. The disclosure provides engineered viable bacteria. In particular, the disclosure provides viable bacteria with mutated outer membrane biosynthetic pathway leading to disruption of the pathway, preferably substantially lacking lipopolysaccharide (LPS, endotoxin) within the outer membrane. The disclosure further provides methods of generating viable bacteria and uses thereof. The disclosure also provides compositions and methods for inducing immune responses and for researching and developing therapeutic agents. Furthermore, the disclosure is in the technical field of fermentation of metabolically engineered microorganisms producing bioproduct or metabolite.

The present invention discloses a transaminase mutant and application thereof in preparation of sitagliptin intermediates, the transaminase mutant is obtained by substitution of tyrosine with proline at position 74, substitution of glutamic acid with aspartic acid at position 228, substitution of leucine with alanine at position 254 and substitution of methionine with threonine at position 290 of the amino acid sequence shown in SEQ ID NO: 2. The present invention uses wet cells or a purified transaminase as a biocatalyst and a sitagliptin precursor ketone or a prochiral carbonyl compound as a substrate to prepare a sitagliptin intermediate or a sitagliptin ester intermediate; the total yield of the method reaches about 82%, and e.e. value of the product reaches 99%.

Methods and compositions are provided for modifying the genome of Bacillus sp. cells without the use of a selectable marker and without the use of a guided Cas endonuclease. The disclosure includes methods for integrating donor DNA sequences into the genome of a Bacillus sp. cell without the use of a selectable marker and without the use of Cas endonucleases into said genome, as well as methods for deleting genes of interest and/or providing point mutations into the genome of Bacillus sp. cells.

The present invention provides a recombinant cell for producing para-nitro-L-phenylalanine (pN-Phe). The recombinant cell comprises heterologous genes encoding heterologous enzymes. The recombinant cell expresses the heterologous enzymes and contains a native metabolite. The native metabolite is converted to the pN-Phe in the recombinant cell. The biosynthesized pN-Phe may be incorporated into a target polypeptide in the recombinant cell without requiring exposure of the recombinant cell to exogenous pN-Phe. A cell culture comprising the recombinant cell is also provided. Further provided is a method of producing pN-Phe by a recombinant cell comprising heterologous genes encoding heterologous enzymes. The method comprises expressing a native metabolite by the recombinant cell, expressing the heterologous enzymes, and converting the native metabolite to the pN-Phe in the recombinant cell. The method may further comprise incorporating the pN-Phe into the target polypeptide in the recombinant cell.

The invention discloses a recombinant Corynebacterium glutamicum for efficient synthesis of highly pure hyaluronic acid and oligosaccharides thereof, belonging to the technical field of bioengineering. The recombinant Corynebacterium glutamicum constructed in the present invention can produce hyaluronic acid with a yield up to 40g/L, and a crude product purity of 95%. Addition of exogenous hyaluronic acid hydrolase and optimization of the fermentation conditions results in hyaluronic acid oligosaccharides with specific molecular weight, and can further improve the yield of hyaluronic acid to 72 g/L. The invention lays a solid foundation for the efficient synthesis of highly pure hyaluronic acid by microorganisms, and the constructed recombinant Corynebacterium glutamicum is suitable for industrial production and application.