EndoH

EndoH, or Endoglycosidase H, is an enzyme that belongs to the class of glycoside hydrolases, specifically those that cleave N-glycans from glycoproteins. It is derived from the bacterium Streptomyces plicatus and is widely used in molecular biology and biochemistry for the analysis and modification of glycoproteins.

EndoH recognizes high-mannose and hybrid-type N-glycans and cleaves them between the chitobiose core and the first mannose residue. This specificity allows it to selectively remove certain glycan structures from glycoproteins, which can be useful for studying glycosylation patterns or simplifying complex glycoproteins for structural or functional analysis. Unlike other endoglycosidases, such as PNGase F, EndoH does not release the entire glycan but instead generates smaller fragments, leaving a single N-acetylglucosamine (GlcNAc) residue attached to the asparagine residue of the original glycosylation site.

In laboratory settings, EndoH is commonly employed in techniques such as Western blotting to assess the glycosylation state of proteins. For example, it can differentiate between endoplasmic reticulum (ER)-type glycoproteins, which contain high-mannose glycans sensitive to EndoH, and Golgi-processed glycoproteins, which often contain complex or hybrid glycans that are resistant to cleavage. This distinction is particularly useful in studying protein trafficking, folding, and quality control within the secretory pathway.

EndoH is typically provided as a recombinant enzyme or in crude extracts and requires specific buffer conditions, such as low pH (around 5.5) and the presence of divalent cations like Mn²⁺ or Co²⁺, for optimal activity. While generally stable under storage conditions, its activity can be influenced by factors such as temperature, pH, and the presence of inhibitors or denaturants. Proper handling and storage, often at −20°C, are recommended to maintain enzymatic function.