Figure 4. Quantitation of glutamine-6 deamidation

Resolution of amidated and deamidated forms of tryptic peptides corresponding to sequence 1-11 of [alpha]-A crystallin. The tryptic digest was resolved on a C₁₈ reverse phase column (4.6 mm x 250 mm, Vydac) using a linear gradient of 23 - 30% (v/v) acetonitrile in 0.1% (v/v) trifluoroacetic acid over a period of 60 min, followed by a linear gradient of 30 - 60% (v/v) acetonitrile in 0.1% (v/v) trifluoroacetic acid, over a period of 5 min. Only the part of the elution profile showing peptides of interest is shown. Top, digest from 71 yr old normal lens (71N); middle, digest from 65 yr old cataractous lens (65C) with opacification of the nucleus; bottom, synthetic peptide standards (STND). Large open arrows designate elution of amidated peptide AcMDVTIQHPWFK, small open arrows designate elution of amidated peptide AcM[0]DVTIQHPWFK, large closed arrows designate elution of deamidated peptide AcMDVTIEHPWFK, small closed arrows designate elution of deamidated peptide AcM[0]DVTIEHPWFK.