The readily available, highly selective (S)-hydroxynitrile lyase from Hevea brasiliensis is used for the synthesis of (S)-cyanohydrins on laboratory and industrial scale. Numerous studies on the deactivation of this important enzyme under a large variety of conditions have been published. Different buffers and their concentrations significantly influence the stability of HbHNL and under acidic conditions, the preferred reaction conditions, it is deactivated particularly fast. However, little attention has so far been paid to the reasons and the mechanism of this deactivation.

Electrospray ionization mass spectrometry (ESI-MS) is a uniquely powerful approach for performing folding analysis on very small quantities within a very short time. This gentle form of ionization in mass spectrometry is ideal for distinguishing between folded and unfolded proteins. We examined the acid induced, irreversible deactivation of HbHNL by ESI-MS, circular dichroism and by an enzyme assay. The deactivation is paralleled by an unfolding of the enzyme. ESI-MS of this 30 000 Da heavy protein demonstrated that unfolding took place in several stages parallel with the deactivation. This unfolding can also be seen by CD. It can be concluded that there is a clear correlation between enzyme activity and unfolding as detected by ESI-MS and CD [1].

[1] U. Hanefeld, G. Stranzl, A.J.J. Straathof, J.J. Heijnen and C. Kratky, Electrospray mass spectrometry and circular dichroism studies of the (S)-hydroxynitrile lyase from Hevea brasiliensis, in preparation.