Recombinant therapeutic glycine n-acyltransferase

Abstract

This invention relates to a method of producing a recombinant enzyme, more particularly, this invention relates to a method of producing water soluble enzymatically active recombinant glycine N-acyltransferase (GLYAT (E.G. 2.1.3.13)), including the steps of providing a suitable expression host; preparing a vector including a gene for expressing GLYAT in the expression host to form an expression piasmid; transforming the host with the expression piasmid to form an expression system; expressing the GLYAT gene in the expression system; and separating the expressed GLYAT from the expression system.

1 . A method of producing water soluble enzymatically active recombinant glycine N-acyltransferase (GLYAT) including the steps of:
 providing a suitable expression host;   preparing a vector including a gene for expressing GLYAT in the expression host to form an expression plasmid;   transforming the host with the expression plasmid to form an expression system;   expressing the GLYAT gene in the expression system; and   separating the expressed GLYAT from the expression system.   
     
     
         2 . A method according to  claim 1  wherein the step of separating the expressed GLYAT from the expression system includes the steps of separating the water soluble fraction of the expression system from the insoluble material and concentrating or lyophilising the separated GLYAT. 
     
     
         3 . A method according to  claim 1  wherein the expression host is selected from the group consisting of eukaryotic systems, including yeast cell expression-, insect cell expression-, and mammalian cell expression systems; prokaryotic systems, including  Escherichia coli , and  Bacillus subtilis ; and archaeon systems. 
     
     
         4 . A method according to  claim 1  including the further step of combining the separated expressed GLYAT with glycine. 
     
     
         5 . Water soluble enzymatically active recombinant GLYAT prepared in accordance with the method of  claim 1 . 
     
     
         6 . A medicament comprising water soluble enzymatically active recombinant GLYAT prepared in accordance with the method of  claim 1   4 . 
     
     
         7 . Use of a pharmaceutically effective amount of water soluble enzymatically active recombinant GLYAT prepared in accordance with the method of  claim 1  in a method of:
 improving the capacity of a glycine-conjugation detoxification system; 
 enhancing detoxification; or 
 treating and/or preventing metabolic disorders and acute or chronic poisoning with compounds such as xylene or aspirin in mammals. 
 
     
     
         8 . Water soluble enzymatically active recombinant GLYAT for use in a method of:
 improving the capacity of a glycine-conjugation detoxification system;   enhancing detoxification; or   treating and/or preventing metabolic disorders and acute or chronic poisoning with compounds such as xylene or aspirin,   
       in mammals by administering to a mammal in need thereof a biologically effective amount of between 0.1 mg and 160 mg of water soluble enzymatically active recombinant GLYAT per kilogram of body weight. 
     
     
         9 . Use of a pharmaceutically effective amount of water soluble enzymatically active recombinant GLYAT prepared in accordance with the method of  claim 1  in a method of manufacturing a medicament for use in:
 improving the capacity of a glycine-conjugation detoxification system; 
 enhancing detoxification; or 
 treating and/or preventing metabolic disorders and acute or chronic poisoning with compounds such as xylene or aspirin in mammals. 
 
     
     
         10 . Use according to  claim 7  wherein the metabolic disorders are any one or more of the conditions selected from the group consisting of organic acidemias selected from propionic acidemia, isovaleric acidemia and glutaric aciduria; aminoacidurias selected from maple syrup urine disease and hyperglycinemia, and urea cycle disorder. 
     
     
         11 . Use according to  claim 7 , wherein the water soluble enzymatically active recombinant GLYAT prepared in accordance with the method of  claim 1  is administered in combination with glycine in order to further stimulate glycine conjugation capacity. 
     
     
         12 . A medicament comprising water soluble enzymatically active recombinant GLYAT prepared in accordance with the method of  claim 1   4  in combination with glycine, together with at least one inert pharmaceutically acceptable carrier or diluents in a dosage form selected from a group comprising tablet; capsule; suspension; syrup; TAT (transactivator of transcription) peptide; a colloidal system that contains unique and stable lipid-based submicron- and micron-sized structures; intradermal-; intramuscular-; intravenous-; and subcutaneous injection. 
     
     
         13 . A method of producing water soluble enzymatically active recombinant glycine N-acyltransferase (GLYAT) as herein described and exemplified, with reference to the accompanying figures.