Provided herein are combination therapies involving co-expression of an E2 subunit of a branched-chain alpha-keto acid dehydrogenase (BCKDH) from a skeletal muscle-targeted rAAV.hDBT vector and a liver-targeted rAAV.hDBT vector. Also provided herein are combination therapies wherein an E1a and/or an E1b subunit of the BCKDH complex is expressed from muscle and/or liver following rAAV-mediated delivery targeted to these tissues. Further provided is a pharmaceutical composition comprising a rAAV as described herein in a formulation buffer, and a method of treating a human subject diagnosed with MSUD.

Genetically modified microorganisms that have the ability to convert carbon substrates into chemical products such as isobutanol are disclosed. For example, genetically modified methanotrophs that are capable of generating isobutanol at high titers from a methane source are disclosed. Methods of making these genetically modified microorganisms and methods of using them are also disclosed.

Maple syrup urine disease (MSUD) is a rare autosomal recessive disease with an incidence that is caused by a defective activity of the branched-chain 2-keto acid dehydrogenase (BCKD) leading to accumulation of branched-chain amino acids (BCAA) leucine, isoleucine, valine and their corresponding alpha-ketoacids (BCKA) in tissues and body fluids. The inventors herein characterized the Bckdha.sup.−/− mouse and Bckdhb.sup.−/− mouse recapitulating the classical forms of MSUD. As a proof of concept, they developed a (liver-directed) AAV gene therapy based on the transfer of human BCKDHA (hBCKDHA) or BCKDHB (hBCKDHB) mediated by AAV8 during immediate neonatal period in Bckdha−/− or Bckdhb.sup.−/− mice. The inventors demonstrated that hBCKDHA gene transfer completely rescued the lethal early-onset phenotype of Bckdha−/− mice allowing long-term survival to age 12 months, at which they were systematically sacrificed, without overt phenotypic abnormalities. They also demonstrated that hBCKDHB gene transfer exhibited similar survival and a normal growth without overt phenotypic abnormalities at age 3 months, with a dramatic improvement of the biochemical phenotype. The present invention relates to a method of treating MSUD by gene therapy.

The present disclosure discloses a genetically engineered strain, belonging to the technical field of bioengineering. L-amino acid oxidase genes, α-keto acid decarboxylase genes, alcohol dehydrogenase genes, and enzyme genes capable of reducing NAD(P) to NAD(P)H are introduced into the genetically engineered strain of the present disclosure. The present disclosure further discloses a construction method and application of a recombinant Escherichia coli genetically engineered strain. When being applied to the biosynthesis of phenylethanoids, the method of the present disclosure has the characteristics of simple operation, low cost, and high synthesis efficiency and optical purity of the product, and has good industrialization prospects.

In some aspects the disclosure provides compositions and methods for promoting expression of functional BCKDHA protein, which is the E1-alpha subunit of the branched-chain alpha-keto acid (BCAA) dehydrogenase complex, in a subject. In some aspects the disclosure provides compositions and methods for promoting expression of functional BCKDHB protein, which is the E1-beta subunit of the branched-chain alpha-keto acid (BCAA) dehydrogenase complex, in a subject. In some aspects the disclosure provides compositions and methods for promoting expression of functional BCKDHA and BCKDHB proteins, in a subject. In some embodiments, the disclosure provides methods of treating a subject having Maple Syrup Urine Disease (MSUD).

Genetically modified microorganisms that have the ability to convert carbon substrates into multicarbon products. Methods of making these genetically modified microorganisms and methods of using them. Vectors encoding enzymes for use in converting carbon substrates into multicarbon products.

This disclosure relates to mRNA therapy for the treatment of maple syrup urine disease (MSUD). mRNAs for use in the invention, when administered in vivo, encode branched chain α-ketoacid dehydrogenase complex (BCKDC) E1α, E1β, or E2mRNA therapies of the disclosure increase and/or restore deficient levels of E1α, E1β, or E2 expression and/or BCKDC activity in subjects. mRNA therapies of the invention further decrease abnormal accumulation of branched chain amino acids associated with deficient BCKDC activity in subjects.

Genetically modified microorganisms that have the ability to convert carbon substrates into chemical products such as isobutanol are disclosed. For example, genetically modified methanotrophs that are capable of generating isobutanol at high titers from a methane source are disclosed. Methods of making these genetically modified microorganisms and methods of using them are also disclosed.

The present invention relates to a process for the production of methyl methacrylate. The process of the present invention comprises the steps of: a) providing a microorganism in a fermentation medium, under conditions which said microorganism will produce a C.sub.3-C.sub.12 methacrylate ester; b) providing an organic phase in contact with the fermentation medium, said organic phase including C.sub.3-C.sub.12 methacrylate ester in a higher concentration than that in the fermentation medium; c) removing organic phase containing the said C.sub.3-C.sub.12 methacrylate ester from contact with the fermentation medium; and d) transesterifying the removed C.sub.3-C.sub.12 methacrylate ester with methanol, optionally after separation from the organic phase, to produce methyl methacrylate.

The present invention relates to a process for the production of methyl methacrylate. The process of the present invention comprises the steps of: a) providing a microorganism in a fermentation medium, under conditions which said microorganism will produce a C.sub.3-C.sub.12 methacrylate ester; b) providing an organic phase in contact with the fermentation medium, said organic phase including C.sub.3-C.sub.12 methacrylate ester in a higher concentration than that in the fermentation medium; c) removing organic phase containing the said C.sub.3-C.sub.12 methacrylate ester from contact with the fermentation medium; and d) transesterifying the removed C.sub.3-C.sub.12 methacrylate ester with methanol, optionally after separation from the organic phase, to produce methyl methacrylate.