US 2019/0194707 A1 · BIOTECHNOLOGICAL METHODS FOR PROVIDING 3,4-DIHYDROXYPHENYL COMPOUNDS AND METHYLATED VARIANTS THEREOF

Abstract

The present invention relates to genetically modified enzymes obtained by rational design of the active site binding pocket of the prototypic enzyme 4-hydroxyphenylacetate 3-hydroxylase (4HPA3H) for hydroxylating a 4-hydroxyphenyl compound to yield a 3,4-dihydroxyphenyl compound and to biotechnological methods including in vivo and in vitro methods using said enzymes or catalytically active fragments thereof. Further provided is a method either using a suitable oxidase or hydroxylase further enabling the subsequent site specific methylation of the 3,4-dihydroxyphenyl compound in a coupled enzymatic reaction by providing a suitable O-methyltransferase. Finally, compositions obtainable by the aforementioned methods are disclosed.

Claims20

Dependent claims

A method for catalyzing the biotechnological conversion of at least one 4-hydroxyphenyl compound for producing at least one 3,4-dihydroxyphenol compound comprising:

(i) providing at least one amino acid sequence comprising a genetically modified Escherichia coli derived 4-hydroxyphenylacetate 3-hydroxylase or a catalytically active fragment thereof, comprising at least one mutation in its active site;
(ii) providing at least one 4-hydroxyphenyl compound;
(iii) reacting the at least one 4-hydroxyphenyl compound and the at least one amino acid sequence comprising the 4-hydroxyphenylacetate 3-hydroxylase or the catalytically active fragment thereof under suitable reaction conditions for allowing the hydroxylation of the at least one 4-hydroxyphenyl compound by the at least one amino acid sequence comprising the 4-hydroxyphenylacetate 3-hydroxylase or the catalytically active fragment thereof to yield at least one 3,4-dihydroxyphenol compound;
and
(iv) optionally isolating and/or purifying the resulting at least one 3,4-dihydroxyphenol compound.

2 more elements

Dependencies are inferred from the claim text.

Biotechnological methods for providing 3,4-dihydroxyphenyl compounds and methylated variants thereof

Life of this patent

Biotechnological methods for providing 3,4-dihydroxyphenyl compounds and methylated variants thereof

Life of this patent

Abstract

Claims20