Strategy for the mass spectrometric verification and correction of the primary structures of proteins deduced from their DNA sequences.

Abstract

Fast atom bombardment mass spectrometry is used to confirm and correct regions from the amino acid sequences of 3 large proteins, glutaminyl- and glycyl-tRNA synthetase from Escherichia coli and methionyl-tRNA synthetase from yeast, whose primary structures were deduced from the base sequences of their corresponding genes. The strategy is based on a comparison of the MW of the tryptic peptides predicted from all 3 reading frames of the gene sequences with those determined mass spectrometrically. The experimental MW match or differ and can be used to assess the correctness of the base sequences, identify errors that lead to frame shifts, premature stop codons, incorrect amino acids, etc., or identify the presence of posttranslational modifications. This method is very fast and requires little material (5-20 nmol).