Nucleic acid molecules encoding clostridium histolyticum collagenase I and methods of producing the same
The invention relates to recombinant nucleic acid and polypeptides encoding collagenase I and collagenase II, methods for the preparation thereof and methods for the use thereof. The invention also encompasses methods related to releasing a composition comprising collagenase prior to therapeutic administration.
1. A recombinant nucleic acid molecule, comprising:
a polynucleotide encoding the amino acid sequence of SEQ ID NO: 5; and
a heterologous regulatory sequence operably linked to the polynucleotide.
2. The recombinant nucleic acid molecule of claim 1 , wherein the heterologous regulatory sequence is a promoter.
3. The recombinant nucleic acid molecule of claim 1 , wherein the recombinant nucleic acid molecule is fused to a marker sequence.
4. The recombinant nucleic acid molecule of claim 3 , wherein the marker sequence encodes a polypeptide selected from a glutathione-S-transferase (GST) fusion protein, a hemagglutinin A (HA) polypeptide from influenza, and a hexa-histidine peptide.
5. A vector comprising the recombinant nucleic acid molecule of claim 1 .
6. The vector of claim 5 , wherein the vector is a plasmid.
7. A recombinant host cell comprising the vector of claim 5 .
8. The recombinant host cell of claim 7 , wherein the host cell is selected from a bacterial cell, a fungal cell, an insect cell, a plant cell, and a mammalian cell.
9. The recombinant host cell of claim 8 , wherein the host cell is E. coli , a Streptomyces species, a Pseudomonas species, Serratia marcescens, Salmonella typhimurium , a yeast cell, a plant cell, a thymocyte, a Chinese hamster ovary cell (CHO), a COS cell, or Lactococcus lactis.
10. A method of producing collagenase I, the method comprising culturing the recombinant host cell of claim 7 under conditions suitable for expression of the polynucleotide and recovering the collagenase I.