Draw the synthesis of an N-glycosylated, integral membrane protein with 3 transm
ID: 95056 • Letter: D
Question
Draw the synthesis of an N-glycosylated, integral membrane protein with 3 transmembrane regions, destined for the plasma membrane. (Start with mRNA in the nucleus and end with the protein in the plasma membrane. You do not have to describe all the glycosylation steps, just what is happening in general and where.) In your diagram, be sure to explain with words or show why the carbohydrate groups of membrane glycoproteins always found on the outer surface of the plasma membrane? Think protein expression, not protein function. Be sure to include the different types of protein modifications in the different organelles.
How would this differ for DNA polyermerase?
How would this differ for a soluble protein that functions in the ER that is also methylated?
Explanation / Answer
The biosynthesis of N-linked glycans occurs through three major steps:-
a. Synthesis of dolichol-linked precursor oligosaccharide
b. En bloc transfer of precursor oligosaccharide to protein
c. Processing of the oligosaccharide
In spite of the diversity all N-glycans are synthesized through a common pathway. A large preformed oligosaccharide is transferred from the lipid dolichol to the side chain of amino acid asparagine and there is covalent linkage to the nitrogen of asparagine side chain. Therefore this process is termed as N-linked glycosylation.
Synthesis of dolichol-linked precursor oligosaccharide:- N-linked glycoprotein synthesis requires a lipid intermediate; dolichol phosphate in contrast to the step-wise addition of sugar groups to the O-linked class of glycoproteins.
Dolichols are polyprenoids containing 17 to 21 isoprene units in which terminal unit is saturated.
En bloc transfer of precursor oligosaccharide to protein:- Synthesis of LLO begins on the cytoplasmic side of the ER membrane through the attachment of GlcNAc to dolichol phosphate.
After the formation of Man5GlcNAc2-PP-Dol, the complex is flipped to the luminal side of the ER membrane where further glycosyltransferase activities complete the synthesis of the en bloc oligosaccharide.
On the cytoplasmic side of the ER membrane, Man and Glc residues are added from dolichol-activated sugars, Dol-P-Man and Dol-P-Glc respectively.
After the synthesis of the LLO, the carbohydrate complex is transferred to an asparagine (Asn, N) residue in a substrate protein through the action of the oligosaccharyltransferase complex (OST). The catalytic subunits of OST are encoded by the STT3A and STT3B genes and accessory subunits are encoded by OST48, OST4, RPN1, RPN2, OST2 and N33 genes.
Processing of the oligosaccharide:- The processing and alteration of the composition of the oligosaccharide ensues and continues as the protein passes through the ER and then into and through the Golgi apparatus. The terminal glucose is removed through the action of processing alpha glucosidase I or GluI a membrane bound enzyme recognizing alpha-1,2-linked glucose.
After the removal of the glucose residues, alpha-mannosidases removes several mannose residues as the protein progresses to the Golgi. The action of various glucosidases and mannosidases leaves N-linked glycoproteins containing a common core of carbohydrate attached to the polypeptide to the polypeptide.
This core consists of three mannose residues and two GlcNAc. A variety of other sugars are attached to this core resulting in the generation of three N-linked glycoprotein families:-
1. High mannose type containing all mannose outside the core in varying amounts.
2. Hybrid type containing various sugars and amino sugars.
3. Complex type which is similar to the hybrid type and in addition contains sialic acids to varying degrees.
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