A rapid and direct method for the determination of active site accessibility in proteins based on ESI-MS and active site titrations

We have developed an electrospray ionisation mass spectrometry (ESI-MS) technique that can be applied to rapidly determine the number of intact active sites in proteins. The methodology relies on inhibiting the protein with an active-site irreversible inhibitor and then using ESI-MS to determine the extent of inhibition. We have applied this methodology to a test system: a serine protease, subtilisin Carlsberg, and monitored the extent of inhibition by phenylmethylsulfonyl fluoride (PMSF), an irreversible serine hydrolase inhibitor as a function of the changes in immobilisation and hydration conditions. Two types of enzyme preparation were investigated, lyophilised enzymes and protein-coated microcrystal

Evaporation and deposition of alkyl-capped silicon nanocrystals in ultrahigh vacuum

Nanocrystals are under active investigation because of their interesting size- dependent properties(1,2) and potential applications(3-5). Silicon nanocrystals have been studied for possible uses in optoelectronics(6), and may be relevant to the understanding of natural processes such as lightning strikes(7). Gas-phase methods can be used to prepare nanocrystals, and mass spectrometric techniques have been used to analyse Au-8,(9) and CdSe clusters(10). However, it is difficult to study nanocrystals by such methods unless they are synthesized in the gas phase(11). In particular, pre-prepared nanocrystals are generally difficult to sublime without decomposition. Here we report the observation that films of alkyl-capped silicon nanocrystals evaporate upon heating in ultrahigh vacuum at 200 degrees C, and the vapour of intact nanocrystals can be collected on a variety of solid substrates. This effect may be useful for the controlled preparation of new quantum-confined silicon structures and could facilitate their mass spectroscopic study and size- selection(12)