Proteins are biomolecules essential for life. Most proteins need to be altered after being synthesized, a process named post-translational modifications. One of the most important post-translational modification is N-glycosylation, in which a sugar molecule becomes attached to an asparagine residue (amino acid) in the protein. Other sugars can subsequently attach to the initial one, forming complex branches that are crucial for the stability of the protein and its correct functioning. Many N-glycosylated proteins, such as the spike protein of SARS- CoV-2, are therapeutic targets.
N-glycosylation is usually initiated by an oligosaccharyltransferase enzyme (OST), that binds a specific oligosaccharide (a short sugar chain) to specific asparagine residues in proteins. It was recently discovered that a bacterial enzyme, identified as soluble N-glycosyltransferase (NGT), could perform a simplified form of N-glycosylation using a single sugar molecule, which could have biotechnological applications (e.g. optimization of the glycan decoration of pharmaceuticals).
In this study the researchers discovered that AaNGT uses a particular pair of basic/acidic residues to recognize the amino acid sequence where the crucial asparagine is located. Furthermore, they used QM/MM MD simulations to uncover the enzyme catalytic mechanism. In this mechanism, the asparagine residue needs to be in its imidic form to react, and the donor substrate itself, rather than an enzyme residue as in other glycosyltransferase enzymes, acts as the general base. These findings are not only significant for enhancing our understanding of NGTs but also open up novel mechanistic pathways for achieving glycosylation that diverges from the most established mechaisms in glycosyltransferases. Additionally, the knowledge inferred from this work might serve to engineer these enzymes to use them for biotechnological applications such as the synthesis of customed N-glycans in molecules as important as antibodies.
