Recombinant lectin and uses thereof

Abstract

Disclosed herein are a recombinant Streptomyces S27S5 hemagglutinin (SHA), and homologues thereof, and a fusion protein of a fluorescent protein (such as GFP and mCherry1) and SHA or a homologue thereof, which specifically bind to carbohydrates, including oligomeric sugars that terminate in L-rhamnose or D-galactose. The SHA, SHA homologues, and fusion proteins can be used to detect a variety of microorganisms or cancer or tumor antigens.

1 . A recombinant  Streptomyces  S27S5 hemagglutinin (SHA) protein having an amino acid sequence represented by SEQ ID NO: 25 or a homologue thereof having an amino acid sequence at least 70%, at least 75%, at least 80%, at least 85%, at least 90%, at least 95%, at least 98%, or at least 99% identical to that of SEQ ID NO: 25, wherein the SHA protein or a homologue thereof specifically binds to L-rhamnose or D-galactose. 
     
     
         2 . The recombinant SHA protein or a homologue thereof of  claim 1 , wherein the recombinant SHA protein or a homologue thereof specifically binds to a carbohydrate that contains or terminates in a monosaccharide or an oligosaccharide selected from the group consisting of β-Gal-; α-Rha-; Gal-α-1,3-Gal-β-1,3-GlcNAc-β-; Gal-α-1,3-Gal-β-1,4-Glc-β-; Gal-α-1,4-Gal-β-1,3-GlcNAc-β-; Gal-α-1,3-(Fuc-α-1,2)-Gal-β-; Gal-α-1,3-(Fuc-α-1,2)-Gal-β-1,4-Glc-β-; Gal-α-1,3-Gal-β-, Gal-α-1,4-Gal-β-1,4-Glc-β-; GalNAc-β-1,3-Gal-β-1,4-Glc-β-; and Gal-α-1,4-Gal-β-1,4-GlcNAc-β-. 
     
     
         3 . A fusion protein comprising a fluorescent protein and SHA, a homologue thereof, or a functional fragment of the SHA or a homologue thereof, wherein the SHA protein has an amino acid sequence represented by SEQ ID NO: 25, the homologue has an amino acid sequence at least 70%, at least 75%, at least 80%, at least 85%, at least 90%, at least 95%, at least 98%, or at least 99% identical to that of SEQ ID NO: 25, and wherein the fusion protein specifically binds to L-rhamnose or D-galactose. 
     
     
         4 . The fusion protein of  claim 3 , wherein the fusion protein specifically binds to a carbohydrate that contains or terminates in a monosaccharide or an oligosaccharide selected from the group consisting of β-Gal-; α-Rha-; Gal-α-1,3-Gal-β-1,3-GlcNAc-β-; Gal-α-1,3-Gal-β-1,4-Glc-β-; Gal-α-1,4-Gal-β-1,3-GlcNAc-β-; Gal-α-1,3-(Fuc-α-1,2)-Gal-β-; Gal-α-1,3-(Fuc-α-1,2)-Gal-β-1,4-Glc-β-; Gal-α-1,3-Gal-β-, Gal-α-1,4-Gal-β-1,4-Glc-β-; GalNAc-β-1,3-Gal-β-1,4-Glc-β-; and Gal-α-1,4-Gal-β-1,4-GlcNAc-β-. 
     
     
         5 . The fusion protein of  claim 3 , wherein the fluorescent protein includes GFP and mCherry1. 
     
     
         6 . A method of detecting a microbial infection caused by a microorganism expressing a carbohydrate containing L-rhamnose or D-galactose on the surface in a subject, comprising:
 contacting a sample obtained from the subject with the fusion protein of  claim 3 ,   detecting the fluorescence level in the sample, wherein the presence of fluorescence indicating a microbial infection.   
     
     
         7 . The method of  claim 6 , wherein the sample includes a biopsy sample, a tissue sample, a bronchoalveolar lavage sample, a blood sample, and a urine sample. 
     
     
         8 . The method of  claim 6 , wherein the microbial infection is caused by a bacterium or a fungus that expresses dTDP-4-dehydrorhamnose reductase gene (rmID). 
     
     
         9 . The method of  claim 6 , wherein the microbial infection includes mycoses caused by  Candida albicans, Aspergillus fumigatus , or  Fusarium solani.    
     
     
         10 . The method of  claim 6 , wherein the microbial infection is an infection by  Streptococcus, Enterococcus  or  Lactococcus.    
     
     
         11 . The method of  claim 6 , wherein the microbial infection is invasive pulmonary aspergillosis, and the GFP-SHA fusion protein detects the presence of fungal galactomannan. 
     
     
         12 . A method of detecting a cancer or tumor in a subject, comprising:
 contacting a sample obtained from the subject with the fusion protein of  claim 3 ,   detecting the fluorescence level in the sample, wherein the presence of fluorescence indicating the presence of the cancer or tumor,   wherein the cancer or tumor cell expresses an antigen comprising a carbohydrate capable of specifically binding to SHA, a homologue thereof, or a fragment of the SHA or a homologue thereof.   
     
     
         13 . The method of  claim 12 , wherein the sample includes a biopsy sample, a tissue sample, a bronchoalveolar lavage sample, a blood sample, and a urine sample. 
     
     
         14 . The method of  claim 12 , wherein the cancer is colon cancer, pancreatic ductal carcinoma, or pancreatic cancer. 
     
     
         15 . The method of  claim 12 , wherein the antigen is a surface antigen comprising a carbohydrate containing D-galactose. 
     
     
         16 . A positron emission tomography (PET) probe comprising the recombinant SHA protein or a homologue thereof of  claim 1  labeled with a PET isotope, or the fusion protein of  claim 3  labeled with a PET isotope. 
     
     
         17 . A method of imaging an organ or tissue having a microbial infection caused by a microorganism expressing a carbohydrate containing L-rhamnose or D-galactose on the surface in a subject, comprising:
 administering to the subject the PET probe of  claim 16 , and   imaging the organ or the tissue having the microbial infection by a PET scanning of the subject.   
     
     
         18 . The method of  claim 17 , wherein the PET probe is administered locally or systemically to the organ or the tissue. 
     
     
         19 . The method of  claim 17 , wherein the PET probe is administered by intravenous injection. 
     
     
         20 . A method of imaging a solid tumor in a subject, comprising:
 administering to the subject the PET probe of  claim 16 , and   imaging the solid tumor by a PET scanning of the subject,   wherein the tumor cell expressing a tumor-specific antigen comprising a carbohydrate capable of specifically binding to SHA, a homologue thereof, or a fragment of the SHA or a homologue thereof.   
     
     
         21 . The method of  claim 20 , wherein the PET probe is administered locally to the tumor. 
     
     
         22 . The method of  claim 20 , wherein the antigen is a surface antigen comprising a carbohydrate containing D-galactose. 
     
     
         23 . A method of detecting the location of a microbial infection caused by a microorganism expressing a carbohydrate containing L-rhamnose or D-galactose on the surface in a subject, comprising:
 administering to the subject the PET probe of  claim 16 , and   detecting the location of the microbial infection by a PET scanning of the subject, wherein the presence of the PET probe indicating the location of the microbial infection.   
     
     
         24 . The method of  claim 23 , wherein the PET probe is administered to the subject by intravenous injection. 
     
     
         25 . A method of detecting the location of cancer cells in a subject, comprising:
 administering to the subject the PET probe of  claim 16 , and   detecting the location of the cancer cells by a PET scanning of the subject, wherein the presence of the PET probe indicating the location of the cancer cells,   and wherein the cancer cells express an antigen comprising a carbohydrate capable of specifically binding to SHA, a homologue thereof, or a fragment of the SHA or a homologue thereof.   
     
     
         26 . The method of  claim 25 , wherein the PET probe is administered to the subject by intravenous injection. 
     
     
         27 . The method of  claim 25 , wherein the antigen is a surface antigen comprising a carbohydrate containing D-galactose.