Abstract
Three human α-amylases exist: Amy1 (salivary amylase), Amy2A (pancreatic amylase), and Amy2B (expressed in various tissues). These amylases share a 97% - 99% amino acid sequence identity, and two potential N-glycosylation sites (N427 and N476) are commonly found in the C-terminal region. In general, salivary amylase is more frequently glycosylated than pancreatic amylase, and it is still uncertain why differences in the glycosylation pattern among human amylase iso-zymes occur. In this study, we found that there was no significant change of ratio of glycosylated molecules among isozymes produced by the same cultured cells, indicating that glycosylation of amylase is influenced by the type of cell producing the enzyme rather than being an inherent property of the amylase isozymes. We analyzed the glycosylation efficiency of N-glycosylation sites in recombinant Amy2A mutants produced by HEK293 cells and found that glycosylation efficiencies of N427 and N476 were 3% - 18% and 40% - 52%, respectively, indicating that the major N-glycosylation site of glycosylated Amy2A produced by HEK293 cells is N476. The difference in the glycosylation efficiency of each N-glycosylation site also seemed to contribute in part to generate different glycosylation patterns of human amylases. We also confirmed that the C-terminal region of human amylase plays a critical role in secretion, although glycosylation does not play a part in this effect.
Highlights
There are three types of human α-amylases: Amy1 is the so-called salivary amylase and is mainly secreted in saliva; Amy2A is the so-called pancreatic amylase and is mainly secreted in pancreatic juice; Amy2B is a pancreatic amylase but it is expressed in various tissues.These amylases share a 97% - 99% amino acid sequence identity and are composed of three domains: the signal peptide, the catalytic domain and the C-terminal region (Figure 1)
In addition to the structural variety of N-glycans, there is a difference in the glycosylation pattern of human amylase isozymes [5,6]: both the N427 and N476 sites in human salivary amylase can be glycosylated to some extent, whereas those in human pancreatic amylase are either glycosylated to a lesser extent or not glycosylated
A small proportion of molecules were fully glycosylated at both the N427 and the N476 sites, about half of the molecules were glycosylated at either of these sites, and the rest of the molecules that showed the same position as Peptide: N-glycosidase (PNGase) F-treated molecule were unglycosylated
Summary
There are three types of human α-amylases: Amy is the so-called salivary amylase and is mainly secreted in saliva; Amy2A is the so-called pancreatic amylase and is mainly secreted in pancreatic juice; Amy2B is a pancreatic amylase but it is expressed in various tissues These amylases share a 97% - 99% amino acid sequence identity and are composed of three domains: the signal peptide, the catalytic domain and the C-terminal region (Figure 1). In the case of human amylase, glycosylation may not be important since the unglycosylated form of amylases is fully secreted and active It is still uncertain, why differences in the glycosylation pattern among human amylase isozymes occur. We discuss the importance of the C-terminal region of human amylase in secretion
Sign-in/Register to view highlights & summary Sign-in/Register to access full text options 
Talk to us
Join us for a 30 min session where you can share your feedback and ask us any queries you have
Schedule a call
