The domains may not be totally modular. Evidence suggests that interactions of SH2 and SH3 domains with each other or with other parts of their parent proteins may be critical for regulation. We wish to compare the dynamics of SH2 and SH3 domains when they are expressed alone versus when they are part of larger proteins. The protein equilibrates for about 100 minutes in an H2O buffer, pH 6.9.
Appropriate controls allow correction for deuterium loss during analysis.
The peptides generated by pepsin are positively identified in other experiments. The peptides are sent into an electrospray mass spectrometer to determine the isotope pattern and average deuterium content of each peptide. Red indicates an increase in exchange while blue indicates a decrease, with the intensity of the color indicative of the amount of change. This may be due to interaction with SH2 or a reduction in the stability of the SH3 b-sheet. The domains do not behave the same when expressed alone as they do in a larger construct.