Galactosylserine in extensin

Area of Science:

• Plant Cell Wall Biology
• Glycobiology
• Protein Chemistry

Background:

• Tomato cell walls contain extensin, a hydroxyproline-rich glycoprotein.
• Extensins are known to be post-translationally modified with carbohydrates.

Purpose of the Study:

• To investigate the glycosylation pattern of extensin in tomato cell walls.
• To identify the specific amino acid residues and linkage types of galactose attachment.

Main Methods:

• Acid hydrolysis of tomato cell walls to remove arabinose substituents.
• Tryptic digestion to isolate glycopeptides.
• Alkaline hydrolysis (NaOH-NaBH4, NaOH-Na2SO3) and hydrazinolysis to analyze glycosidic linkages.
• Chemical modification (maleylation, 3-carboxypropionylation) of N-terminal serine.

Main Results:

• Acid-stripped cell walls yielded glycopeptides containing galactose.
• Alkaline treatment and hydrazinolysis of a specific glycopeptide (S2A6) resulted in the loss of galactose and conversion of serine to alanine or cysteic acid.
• Chemical modification of N-terminal serine was required for complete galactose loss, indicating O-glycosidic linkage to serine residues.
• Hydrazinolysis destroyed non-glycosylated serine but not glycosylated serine, confirming the galactosylserine linkage sensitivity.

Conclusions:

• A single galactose residue is attached O-glycosidically to two serine residues within the extensin glycopeptide S2A6.
• The galactosylserine linkage is sensitive to alkaline conditions and hydrazinolysis.
• This study elucidates a specific glycosylation site and linkage type in plant extensin.