Title:  Hepatitis C E1 and E2 truncated polypeptides and methods of obtaining the same

United States Patent:  6,326,171

Assignee:  Chiron Corporation (Emeryville, CA)

Filed:  October 8, 1999

Novel Hepatitis C E1 and E2 truncated polypeptides and complexes comprising these polypeptides, are disclosed. The polypeptides are C-terminally truncated to remove all or a portion of their membrane spanning domains. Hence, the polypeptides are capable of secretion when expressed recombinantly.

SUMMARY OF THE INVENTION

The present invention is based on the elucidation of sequences of E1 and E2 important for anchoring the proteins to the endoplasmic reticulum (ER) and for co-precipitation of E2 with E1. Thus, the elimination of these sequences serves to produce truncated forms of the glycoproteins which are secreted rather than retained in the ER membrane. Furthermore, truncation facilitates purification of the E2 protein without associated E1, and vice versa. The truncated E1 and E2 proteins, when expressed together or combined after expression, are capable of forming a complex.

Accordingly, in one embodiment, the subject invention is directed to an HCV E1 polypeptide, lacking all or a portion of its membrane spanning domain such that the polypeptide is capable of secretion into growth medium when expressed recombinantly in a host cell. In preferred embodiments, the HCV E1 polypeptide lacks at least a portion of its C-terminus beginning at or near about amino acid 370 or about amino acid 360, numbered with reference to the HCV1 E1 amino acid sequence.

In another embodiment, the invention is directed to an HCV E2 polypeptide lacking at least a portion of its membrane spanning domain such that the polypeptide is capable of secretion into growth medium when expressed recombinantly in a host cell, wherein the polypeptide lacks at least a portion of its C-terminus beginning at or near about amino acid 730 but not extending beyond about amino acid 699, numbered with reference to the HCV1 E2 amino acid sequence. In particularly preferred embodiments, the HCV E2 polypeptide lacks at least a portion of its C-terminus beginning at about amino acid 725, numbered with reference to the HCV1 E2 amino acid sequence.

Other embodiments of the subject invention pertain to polynucleotides encoding the above polypeptides, vectors comprising these polynucleotides, host cells transformed with the vectors and methods of recombinantly producing the polypeptides.

In yet another embodiment, the subject invention is directed to a secreted E1/secreted E2 complex comprising:

(a) an HCV E1 polypeptide, lacking all or a portion of its membrane spanning domain such that the E1 polypeptide is capable of secretion into growth medium when expressed recombinantly in a host cell; and

(b) an HCV E2 polypeptide, lacking all or a portion of its membrane spanning domain such that said E2 polypeptide is capable of secretion into growth medium when expressed recombinantly in a host cell.

In preferred embodiments, the secreted E1/secreted E2 complex includes an E1 polypeptide which lacks at least a portion of its C-terminus beginning at about amino acid 360 or 370, numbered with reference to the HCV1 E1 amino acid sequence, and said E2 polypeptide lacks at least a portion of its C-terminus beginning at about amino acid 725 or 730, numbered with reference to the HCV1 E2 amino acid sequence.

In still further embodiments, the subject invention is directed to vaccine compositions comprising the truncated HCV E1 and/or E2 polypeptides and/or complexes of the E1 and E2 polypeptides.

Claim 1 of 4 Claims

What is claimed is:

1. A method of producing a secreted HCV polypeptide, said method comprising:

(a) providing a population of mammalian or yeast host cells;

(b) transforming said host cells with a recombinant vector comprising (i) a polynucleotide encoding an HCV E2 polypeptide having a C-terminus at about amino acid residue 715, numbered with reference to the HCV1 E2 amino acid sequence and (ii) control elements that are operably linked to said coding sequence whereby said coding sequence can be transcribed and translated in the host cell, and

(c) culturing said host cells under conditions whereby said HCV polypeptide is expressed and secreted.

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