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Production in Escherichia coli of extracellular Serratia spp. hydrolases
| Details |
Inventors: Molin, Soren; Givskov, Michael; Riise, Erik;
Assignee: Benzon Pharma, A/S (Hvidovre, DK)
Primary Examiner: Schwartz; Richard A.
Assistant Examiner: Ketter; James
Attorney, Agent or Firm: Cooper; Iver P.
Extracellular Serratia spp. enzymes have been found to be excreted by another gram-negative organism harbouring a plasmid carrying DNA from Serratia spp. encoding the enzymes. This organism, e.g. E. coli, is therefore employed to produce the enzymes, specific examples of Serratia spp. enzymes produced are a nuclease, a lipase and a phosopholipase. The nuclease may be employed to remove nucleic acids from a biological material. |
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DETAILED DESCRIPTION We claim: 1.
A method of producing an extracellular hydrolytic Serratia spp.
enzyme selected from the group consisting of exocellular nuclease, exocellular phospholipase and exocellular lipase, comprising cultivating, E.
coli in a culture medium, said E.
coli harboring a hybrid plasmid which comprises DNA encoding an extracellular hydrolytic Serratia spp.
enzyme selected from the group consisting of exocellular nuclease, exocellular phospholipase and exocellular lipase, operably linked to a promotor functional in said E.
coli, under conditions conductive to the expression of said enzyme and its secretion into the culture medium, and harvesting the enzyme from the culture medium.
2.
A method according to claim 1 wherein the Serratia spp.
enzyme is a Serratia spp.
nuclease.
3.
A method according to claim 1 wherein the Serratia spp.
enzyme is a Serratia spp.
phospholipase.
4.
The method of claim 1 wherein the hybrid plasmid is a plasmid with a conditionally uncontrolled replication behavior.
5.
The method of claim 1 wherein the DNA sequence is ##STR3## 6.
A method according to claim 1 of producing the enzyme substantially free from other bacterial protein, in which the enzyme is secreted from the E.
coli into the culture medium and is harvested from the culture medium.
7.
The method of claim 1 wherein the DNA encodes a Serratia spp.
nuclease which, prior to removal of the amino-terminal signal peptide, has the amino acid sequence ##STR4## 8.
The method of claim 1 wherein the DNA encodes a Serratia spp.
phospholipase encoded by the DNA sequence ##STR5## 9.
A DNA fragment comprising a DNA sequence encoding a Serratia spp.
phospholipase.
10.
A DNA fragment according to claim 9 wherein the DNA sequence is ##STR6##
Description:
The present invention relates to a method of producing bacterial enzymes and to hybrid plasmids and microorganisms useful in the method.
The invention further relates to the use of one of the enzymes, nuclease, for removing nucleic acids from a biological material, as well as to a regulatory region useful for initiating gene expression
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