Electrospray ionization mass spectrometry, circular dichroism and SAXS studies of the (S)-hydroxynitrile lyase from Hevea brasiliensis.
We report on experiments pertaining to solution properties of the (S)-hydroxynitrile lyase from Hevea brasiliensis (HbHNL). Small angle X-ray scattering unequivocally established the enzyme to occur in solution as a dimer, presumably of the same structure as in the crystal. The acid induced, irreversible deactivation of HbHNL was examined by electrospray ionization mass spectrometry (ESI-MS), circular dichroism (CD) and by measuring the enzyme activity. The deactivation is paralleled by an unfolding of the enzyme. ESI-MS of this 30000 Da per monomer heavy protein demonstrated that unfolding took place in several stages which are paralleled by a decrease in enzyme activity. Unfolding can also be observed by CD spectroscopy, and there is a clear correlation between enzyme activity and unfolding as detected by ESI-MS and CD.
Hanefeld
U
U
Kluyverlaboratorium voor Biotecnologie, Techische Universiteit Delft, The Netherlands. u.hanefeld@tnw.tudelft.nl
Stranzl
G
G
Straathof
A J
AJ
Heijnen
J J
JJ
Bergmann
A
A
Mittelbach
R
R
Glatter
O
O
Kratky
C
C
D000446:Aldehyde-Lyases / Q000037:antagonists & inhibitors / Q000378:metabolism*; D002942:Circular Dichroism; D019665:Euphorbiaceae / Q000201:enzymology*; D006863:Hydrogen-Ion Concentration; D012542:Scattering, Radiation; D021241:Spectrometry, Mass, Electrospray Ionization; D014965:X-Rays
D000446:Aldehyde-Lyases; C079359:hydroxymandelonitrile lyase