Recombinant host for producing l-asparaginase ii

Abstract

The invention provides a recombinant Escherichia coli host cell for producing an Escherichia coli L-asparaginase II enzyme. The host cell includes an Escherichia coli chromosome and at least one copy of a recombinant extrachromosomal vector, wherein the recombinant extrachromosomal vector encodes the L-asparaginase II enzyme, wherein the host cell chromosome also encodes the same L-asparaginase II enzyme, and wherein the host chromosome does not encode any other isoform of L-asparaginase II.

1 . A recombinant  Escherichia coli  host cell for producing an  Escherichia coli  L-asparaginase II enzyme, comprising an  Escherichia coli  host cell chromosome and at least one copy of a recombinant extrachromosomal vector, wherein the recombinant extrachromosomal vector encodes a subunit of the L-asparaginase II enzyme, wherein the host cell chromosome also encodes the same subunit of the L-asparaginase II enzyme, and wherein the host cell chromosome does not encode any other isoform of L-asparaginase II, wherein the encoded L-asparaginase II subunit comprises SEQ ID NO: 1. 
     
     
         2 . The recombinant  Escherichia coli  host cell of  claim 1 , wherein the extrachromosomal vector is a plasmid. 
     
     
         3 . The recombinant  Escherichia coli  host cell of  claim 1  wherein the recombinant extrachromosomal vector comprises a DNA molecule encoding the L-asparaginase protein, that is operatively connected to a suitable promoter. 
     
     
         4 . The recombinant  Escherichia coli  host cell of  claim 3  wherein the promoter is selected from the group consisting of T7, araB, P R /P L , phoA, trc, and trp promoters. 
     
     
         5 . The recombinant  Escherichia coli  host cell of  claim 3  wherein the recombinant extrachromosomal vector further comprises an operator, ribosome binding site, signal sequence, transcriptional terminator, antibiotic selection marker, origin of replication, and a regulated copy of the repressor. 
     
     
         6 . A method of producing a recombinant L-asparaginase II enzyme substantially free of other L-asparaginase II isomers, comprising culturing the host cell of  claim 1 , and isolating the produced L-asparaginase II enzyme.