Polypeptide Having Affinity for Envelope Virus Constituent and Use Thereof in Transferring Substance Into Cell

Abstract

Proteins are provided for transferring a protein, an antibody or another foreign substance into a cell without impairing the function or structure thereof; and methods of transferring a foreign substance into a cell in a time and quantity controllable manner at a high efficiency by using the above-described delivery protein or an envelope virus or inactivated envelope virus in combination with said delivery protein. As the results of intensive studies on a method of enclosing a foreign substance in the envelope of an envelope virus, it is found out that a protein containing a polypeptide having an affinity for a constituent of the envelope virus contributes to the efficient enclosure of the foreign substance in the envelope. Moreover, it is found out that use of the aforementioned protein enables foreign substances to be included in an envelope virus or inactivated envelope virus and therefore the resulting foreign substance-containing envelope virus or inactivated envelope virus makes it possible to efficiently transfer the substances into cells without damaging the physiological function thereof.

1 . A protein comprising (a) a polypeptide having an affinity for one or more envelope viral components in combination with (b) a polypeptide capable of binding to one or more foreign substances to be introduced into a cell. 
     
     
         2 . The protein according to  claim 1 , wherein the protein comprising said polypeptide (a) in combination with the polypeptide (b) is a fusion protein. 
     
     
         3 . The protein according to  claim 1  or  2 , wherein said polypeptide (a) is a polypeptide having an affinity for one or more components existing in the inner space of an envelope. 
     
     
         4 . The protein according to  claim 1  or  2 , wherein said polypeptide (a) is
 (1) a polypeptide of an amino acid sequence of SEQ ID NO: 2,   (2) a polypeptide having an amino acid sequence substantially equivalent to the amino acid sequence of SEQ ID NO: 2, or   (3) a peptide fragment (partial peptide) thereof.   
     
     
         5 . The protein according to  claim 1  or  2 , wherein said polypeptide (b) is an antibody binding polypeptide. 
     
     
         6 . The protein according to  claim 5 , wherein said antibody binding polypeptide is a polypeptide that can bind to an antibody Fc region or kappa (K) light chain. 
     
     
         7 . The protein according to  claim 5 , wherein said antibody binding polypeptide is
 (1) a polypeptide of an amino acid sequence selected from the group consisting of SEQ ID NOs: 4, 49 and 51,   (2) a polypeptide having an amino acid sequence substantially equivalent to the aforementioned amino acid sequence (1), or   (3) a peptide fragment (partial peptide) thereof.   
     
     
         8 . A DNA encoding a fusion protein consisting of a polypeptide having an affinity for one or more envelope viral components and a polypeptide capable of binding to an antibody wherein said fusion protein is a polypeptide selected from the group consisting of:
 (1) a fusion polypeptide consisting of a polypeptide of an amino acid sequence of SEQ ID NO: 2, a polypeptide substantially equivalent to the amino acid sequence of SEQ ID NO: 2, or a peptide fragment thereof, fused with a polypeptide of an amino acid sequence selected from the group consisting of SEQ ID NOs: 4, 49 and 51, a polypeptide having an amino acid sequence substantially equivalent to the aforementioned amino acid sequence of SEQ ID NO: 4, 49 or 51, or a peptide fragment thereof,   (2) a polypeptide of an amino acid sequence selected from the group consisting of SEQ ID NOs: 6, 13, 55 and 59, and   (3) a polypeptide having an amino acid sequence substantially equivalent to a polypeptide of an amino acid sequence selected from the group consisting of SEQ ID NOs: 6, 13, 55 and 59.   
     
     
         9 . An envelope virus or inactivated envelope virus containing a protein comprising (a) a polypeptide having an affinity for one or more envelope viral components in combination with (b) a polypeptide capable of binding to one or more foreign substances to be introduced into a cell. 
     
     
         10 . The envelope virus or inactivated envelope virus according to  claim 9 , wherein said virus contains a fusion protein consisting of a polypeptide having an affinity for one or more envelope viral components and an antibody-binding polypeptide wherein said fusion protein is a polypeptide selected from the group consisting of:
 (1) a fusion polypeptide consisting of a polypeptide of an amino acid sequence of SEQ ID NO: 2, a polypeptide substantially equivalent to the amino acid sequence of SEQ ID NO: 2, or a peptide fragment thereof, fused with a polypeptide of an amino acid sequence selected from the group consisting of SEQ ID NOs: 4, 49 and 51, a polypeptide having an amino acid sequence substantially equivalent to the aforementioned amino acid sequence of SEQ ID NO: 4, 49 or 51, or a peptide fragment thereof,   (2) a polypeptide of an amino acid sequence selected from the group consisting of SEQ ID NOs: 6, 13, 55 and 59, and   (3) a polypeptide having an amino acid sequence substantially equivalent to a polypeptide of an amino acid sequence selected from the group consisting of SEQ ID NOs: 6, 13, 55 and 59.   
     
     
         11 . An envelope virus or inactivated envelope virus containing a complex constituted of (i) a protein comprising (a) a polypeptide having an affinity for one or more envelope viral components in combination with (b) a polypeptide capable of binding to one or more foreign substances to be introduced into a cell, and (ii) one or more foreign substances. 
     
     
         12 . The envelope virus or inactivated envelope virus according to  claim 11 , wherein said foreign substance is an antibody. 
     
     
         13 . The envelope virus or inactivated envelope virus according to  claim 12 , wherein said virus contains a complex constituted of a fusion protein consisting of a polypeptide having an affinity for one or more envelope viral components, fused to an antibody binding polypeptide, and
 an antibody,   
       wherein said fusion protein is a polypeptide selected from the group consisting of:
 (1) a fusion polypeptide consisting of a polypeptide of an amino acid sequence of SEQ ID NO: 2, a polypeptide substantially equivalent to the amino acid sequence of SEQ ID NO: 2, or a peptide fragment thereof, fused with a polypeptide of an amino acid sequence selected from the group consisting of SEQ ID NOs: 4, 49 and 51, a polypeptide having an amino acid sequence substantially equivalent to the aforementioned amino acid sequence of SEQ ID NO: 4, 49 or 51, or a peptide fragment thereof, 
 (2) a polypeptide of an amino acid sequence selected from the group consisting of SEQ ID NOs: 6, 13, 55 and 59, and 
 (3) a polypeptide having an amino acid sequence substantially equivalent to a polypeptide of an amino acid sequence selected from the group consisting of SEQ ID NOs: 6, 13, 55 and 59. 
 
     
     
         14 . A process for preparing an envelope virus or inactivated envelope virus containing a foreign substance within an envelope, which comprises the steps consisting of
 (1) mixing an envelope virus or inactivated envelope virus with a protein comprising (a) a polypeptide having an affinity for one or more envelope viral components in combination with (b) a polypeptide capable of binding to one or more foreign substances to be introduced into a cell,   (2) subjecting the resultant mixture from the above step (1), to a treatment affecting the permeability of envelopes, and   (3) mixing the resultant product from the above step (2), with one or more foreign substances.   
     
     
         15 . A process for preparing an envelope virus or inactivated envelope virus containing a foreign substance within an envelope, which comprises the steps consisting of
 (1) mixing a protein comprising (a) a polypeptide having an affinity for one or more envelope viral components in combination with (b) a polypeptide capable of binding to one or more foreign substances to be introduced into a cell, with one or more foreign substances to form a complex,   (2) mixing the resultant mixture from the above step (1), with an envelope virus or inactivated envelope virus, and   (3) subjecting the resultant mixture from the above step (2), to a treatment affecting the permeability of envelopes.   
     
     
         16 . The process according to  claim 14  or  15 , wherein the protein comprising said polypeptide (a) in combination with the polypeptide (b) is a fusion protein. 
     
     
         17 . The process according to  claim 14  or  15 , wherein said foreign substance is an antibody. 
     
     
         18 . A method for introducing one or more foreign substances into a cell with an envelope virus or inactivated envelope virus, which comprises the steps consisting of
 (1) mixing an envelope virus or inactivated envelope virus with a protein comprising (a) a polypeptide having an affinity for one or more envelope viral components in combination with (b) a polypeptide capable of binding to one or more foreign substances to be introduced into a cell,   (2) subjecting the resultant mixture from the above step (1), to a treatment affecting the permeability of envelopes,   (3) mixing the resultant product from the above step (2) with one or more foreign substances, and   (4) contacting the resulting foreign substance-containing envelope virus or inactivated envelope virus from the above step (3), with one or more cells.   
     
     
         19 . A method for introducing one or more foreign substances into a cell with an envelope virus or inactivated envelope virus, which comprises the steps consisting of
 (1) mixing a protein comprising (a) a polypeptide having an affinity for one or more envelope viral components in combination with (b) a polypeptide capable of binding to one or more foreign substances to be introduced into a cell, with one or more foreign substances to form a complex,   (2) mixing the resultant mixture from the above step (1), with an envelope virus or inactivated envelope virus,   (3) subjecting the resultant mixture from the above step (2), to a treatment affecting the permeability of envelopes and   (4) contacting the resulting foreign substance-containing envelope virus or inactivated envelope virus from the above step (3), with one or more cells.   
     
     
         20 . The process according to  claim 18  or  19 , wherein the protein comprising said polypeptide (a) in combination with the polypeptide (b) is a fusion protein. 
     
     
         21 . The process according to  claim 18  or  19 , wherein said foreign substance is an antibody. 
     
     
         22 . A complex constituted of (i) a protein comprising (a) a polypeptide having an affinity for one or more envelope viral components in combination with (b) a polypeptide capable of binding to one or more foreign substances to be introduced into a cell, and (ii) one or more foreign substances to be introduced into one or more cells.