US 2023/0140642 A1 · MUTATED SULFOTRANSFERASES AND USES THEREOF

Abstract

The invention relates to non-naturally occurring mutated arylsulfotransferases comprising (i) an amino acid substitution in at least one amino acid position selected among positions 6, 7, 8, 9, 11, 17, 20, 33, 62, 97, 138, 195, 236, 239, 244, 263, and combinations thereof, wherein the position is relative to the amino acids sequence of rat arylsulfotransferase IV SEQ ID NO: 1 and (ii) an amino acid sequence having at least 60% sequence identity with amino acids sequence SEQ ID NO: 1. The mutated arylsulfotransferase may have a sulfotransferase activity for converting adenosine 3′,5′-bisphosphate (PAP) into 3′-phosphoadenosine-5′-phosphosulfate (PAPS) enhanced compared to the wild-type enzyme.

Claims19

Dependent claims

A non-naturally occurring mutated arylsulfotransferase comprising (i) an amino acid substitution in at least one amino acid position selected from positions 6, 7, 8, 9, 11, 17, 20, 33, 62, 97, 138, 195, 236, 239, 244, 263, and-combinations thereof, wherein the position is relative to the amino acid sequence of rat arylsulfotransferase IV SEQ ID NO:

1, and (ii) an amino acid sequence having at least 60% sequence identity with amino acid sequence of SEQ ID NO:
1, with the proviso that when said arylsulfotransferase is the rat arylsulfotransferase IV, the mutations are not F138A and/or Y236A.

A method of selecting a non-naturally occurring mutated arylsulfotransferase comprising at least one amino acid substitution and comprising a sulfotransferase activity for converting adenosine 3′,5′-bisphosphate (PAP) into 3′-phosphoadenosine-5′-phosphosulfate (PAPS) being at least 1.3 times greater than the said activity of the rat arylsulfotransferase IV of SEQ ID NO:

1 or being at least substantially the same as or greater than said activity of a non-naturally occurring mutated arylsulfotransferase having an amino acid sequence selected from the group consisting of SEQ ID NO:
5 to 23, 25-35, 41, 45-47, and 49-56, said method comprising the steps of:
a. contacting a non-naturally occurring mutated arylsulfotransferase candidate comprising at least one amino acid substitution with a sulfo group donor in conditions suitable for a transfer of the sulfo group from the sulfo group donor to PAP to obtain PAPS,
b. detecting a rate or an amount of formation of PAPS,
c. comparing the rate or amount of formation of PAPS obtained at step b) with a rate or an amount of reference obtained with a rat arylsulfotransferase IV of SEQ ID NO:
1 or obtained with a non-naturally occurring mutated arylsulfotransferase having an amino acid sequence selected from the group consisting of SEQ ID NO:
5 to 23, 25-35, 41, 45-47, and 49-56, and
d. selecting any non-naturally occurring mutated arylsulfotransferase candidate comprising at least one amino acid substitution and comprising a sulfotransferase activity for converting adenosine 3′,5′-bisphosphate (PAP) into 3′-phosphoadenosine-5′-phosphosulfate (PAPS) being at least 1.3 times greater than the said activity of the rat arylsulfotransferase IV of SEQ ID NO:
1 or being at least substantially the same as said activity of a non-naturally occurring mutated arylsulfotransferase having an amino acid sequence selected from the group consisting of SEQ ID NO:
5 to 23, 25-35, 41, 45-47, and 49-56.

7 more elements

Dependencies are inferred from the claim text.

Mutated sulfotransferases and uses thereof

Life of this patent

Mutated sulfotransferases and uses thereof

Life of this patent

Abstract

Claims19