Protease and Binding Polypeptide for O-Glycoproteins

A method of binding to an O-glycan, O-glycopeptide and/or O-glycoprotein wherein the method comprises contacting a sample comprising the O-glycan, O-glycopeptide and/or O-glycoprotein with a polypeptide which is capable of binding to an O-glycan or O-glycoprotein and which lacks or has reduced endoprotease activity specific for O-glycosylated proteins comprising:

• (a) an amino acid sequence of SEQ ID NO:
• 5 or SEQ ID NO:
• 20;
• (b) an amino acid sequence which is at least 85% identical to the amino acid sequence of SEQ ID NO:
• 5 or SEQ ID NO:
• 20;
• or
• (c) an amino acid sequence which is a fragment of the sequence of SEQ ID NO:
• 5 or SEQ ID NO:
• 20 or a fragment of an amino acid which is 85% identical to the amino acid sequence of SEQ ID NO:
• 5 or SEQ ID NO:
• 20;
• and optionally determining whether or not an O-glycan, O-glycopeptide and/or O-glycoprotein has been bound and/or separating the O-glycan and any linked glycoprotein, the O-glycopeptide or O-glycoprotein from the resulting mixture;
• optionally wherein the method is for the purpose of isolating an O-glycan or linked glycoprotein, O-glycopeptide or O-glycoprotein from the sample.

A method for assessing the glycosylation status of a protein, comprising contacting a sample comprising the protein with a polypeptide which is capable of binding to an O-glycan or O-glycoprotein and which lacks or has reduced endoprotease activity specific for O-glycosylated proteins comprising:

• (a) an amino acid sequence of SEQ ID NO:
• 5 or SEQ ID NO:
• 20;
• (b) an amino acid sequence which is at least 85% identical to the amino acid sequence of SEQ ID NO:
• 5 or SEQ ID NO:
• 20;
• or
• (c) an amino acid sequence which is a fragment of the sequence of SEQ ID NO:
• 5 or SEQ ID NO:
• 20 or a fragment of an amino acid which is 85% identical to the amino acid sequence of SEQ ID NO:
• 5 or SEQ ID NO:
• 20;
• and determining whether or not the protein is bound by the said polypeptide.

A method for detecting O-glycopeptides and/or O-glycoproteins in a sample, wherein the method comprises:

• (a) contacting said sample with a polypeptide which is capable of binding to an O-glycan or O-glycoprotein an which lacks or has reduced endoprotease activity specific for O-glycosylated proteins comprising:
• (i) an amino acid sequence of SEQ ID NO:
• 5 or SEQ ID NO:
• 20;
• (ii) an amino acid sequence which is at least 85% identical to the amino acid sequence of SEQ ID NO:
• 5 or SEQ ID NO:
• 20;
• or
• (iii) an amino acid sequence which is a fragment of the sequence of SEQ ID NO:
• 5 or SEQ ID NO:
• 20 or a fragment of an amino acid which is 85% identical to the amino acid sequence of SEQ ID NO:
• 5 or SEQ ID NO:
• 20;
• to thereby allow formation of complex between an O-glycopeptide and/or O-glycoprotein and the said polypeptide;
• (b) optionally separating said polypeptide from the contacted sample;
• and
• (c) determining whether the separated polypeptide is bound to an O-linked glycoprotein or glycopeptide, thereby determining the presence or absence of O-linked glycopeptides or glycoproteins in the sample.