Patent ID: 6017726
Filing Date: 2000-01-25
Classification: C12N,C12P,Y10S

Abstract:
A bioprocess for preparing 7-aminodeacetylcephalosoporanic acid (7-ADAC) comprising the steps of1) maintaining in a culture medium capable of sustaining its growth, a strain of Penicillium chrysogenum which produces isopenicillin N and adding to said culture medium an adipate feedstocks comprising any one or more of adipic acid, or its salts and esters which are capable of being assimilated and utilized by said strain of Penicillium chrysogenum to produce adipoyl-6-aminopenicillanic acid (adipoyl-6-APA), whereby said adipoyl-6-APA is produced;2) carrying out the following enzymatic conversion by in situ expression of the corresponding gene:a) adipoyl-6-APA is in situ ring-expanded to form adipoyl-7-amino-desacetoxycephalosporanic acid (adipoyl-7-ADCA) by expandase enzyme, wherein said strain of P. chrysogenum has been transformed by DNA encoding the expandase enzyme capable of accepting said adipoyl-6-APA as a substrate, whereupon as a result of its expression, said adipoyl-6-APA produced by said strain is also thereafter in situ ring-expanded to form adipoyl-7-ADCA;b) the 3-methyl side chain of adipoyl-7-ADCA is in situ hydroxylated in yield adipoyl-7-ADAC by hydroxylase enzyme, wherein said strain of P. chrysogenum has been transformed by DNA encoding the hydroxylase enzyme capable of accepting said adipoyl-7-ADCA as a substrate, whereupon as a result of its expression, said adipoyl-7-ADCA produced by said strain is also thereafter in situ hydroxylated to form adipoyl-7-ADAC;3) contacting said adipoyl-7-ADAC with an adipoyl amidase whereby the adipoyl side chain is removed and the 7-ADAC product is formed; and said product is then isolated.