1 research outputs found
Empirical Correction for Differences in Chemical Exchange Rates in Hydrogen Exchange-Mass Spectrometry Measurements
A barrier
to the use of hydrogen exchange-mass spectrometry (HX-MS)
in many contexts, especially analytical characterization of various
protein therapeutic candidates, is that differences in temperature,
pH, ionic strength, buffering agent, or other additives can alter
chemical exchange rates, making HX data gathered under differing solution
conditions difficult to compare. Here, we present data demonstrating
that HX chemical exchange rates can be substantially altered not only
by the well-established variables of temperature and pH but also by
additives including arginine, guanidine, methionine, and thiocyanate.
To compensate for these additive effects, we have developed an empirical
method to correct the hydrogen-exchange data for these differences.
First, differences in chemical exchange rates are measured by use
of an unstructured reporter peptide, YPI. An empirical chemical exchange
correction factor, determined by use of the HX data from the reporter
peptide, is then applied to the HX measurements obtained from a protein
of interest under different solution conditions. We demonstrate that
the correction is experimentally sound through simulation and in a
proof-of-concept experiment using unstructured peptides under slow-exchange
conditions (pD 4.5 at ambient temperature). To illustrate its utility,
we applied the correction to HX-MS excipient screening data collected
for a pharmaceutically relevant IgG4 mAb being characterized to determine
the effects of different formulations on backbone dynamics