B-transferase with a Pro234Ser substitution acquires AB-transferase activity

Area of Science:

• Biochemistry
• Immunogenetics
• Molecular Biology

Background:

• A/B-transferase enzymes are crucial for synthesizing A/B-antigens, which are glycoprotein and glycolipid conjugates.
• A Pro234Ser substitution in B-transferase has been linked to the cis-AB blood group phenotype.

Purpose of the Study:

• To investigate the functional consequences of the Pro234Ser mutation in B-transferase on enzymatic specificity.
• To understand the molecular mechanisms underlying the cis-AB phenotype.

Main Methods:

• Generated expression plasmids for mutated B-transferase (Pro234Ser) and A-transferase (Leu266Met).
• Transfected these constructs into HeLa cells (O-blood group).
• Analyzed resulting phenotypes using absorption-elution testing, immunostaining, and molecular simulation.

Main Results:

• HeLa cells expressing B-transferase/P234S or A-transferase/L266M exhibited an AB-phenotype.
• Cells expressing wild-type A- or B-transferase showed only their respective activities.
• Molecular simulation revealed the P234S mutation induces a conformational change in the substrate pocket, favoring N-acetylgalactosamine binding.

Conclusions:

• The Pro234Ser mutation in B-transferase confers enzymatic activity capable of producing AB-antigens.
• This mutation is a potential molecular basis for the cis-AB blood group phenotype.
• Understanding these enzymatic alterations is key to blood group antigen research.