CHAPTER 9
Detection of Specific Zinc Finger Peptide Complexes
with Matrix-Assisted Laser Desorption/Ionization Mass
Spectrometry
E. LEHMANN l and R. ZENOBI l
1
Introduction
Noncovalent complexes play an important role in biochemistry and molecular
biology. Various methods have been used to investigate these complexes. Among
them, size exclusion chromatography and gel electrophoresis, for example, are
used for determining stoichiometry and molecular weights. These conventional
methods are relatively time- and material-consuming. A recent analytical tool to
overcome these problems is mass spectrometry (MS), characterized by high sensitivity and speed. Together with electrospray ionization (ESI) mass spectrometry, matrix-assisted laser desorption/ionization (MALDI) mass spectrometry is a
soft ionization technique, i.e. leads to little fragmentation of the analyte. For this
reason, it is a powerful method for the analysis of various biomolecules and synthetic polymers (Bahr et al. 1992; Hillenkamp et al. 1991). MALDI spectra are
generated by laser irradiation of analyte molecules embedded in an excess of a
crystalline matrix. This matrix is responsible for the absorption of the incident
laser light and also plays an important role in analyte ionization.
A relevant question to ask is whether a correspondence exists between
solution-phase behavior and the observed gas-phase ions and how soft MALDI is
for the analysis of specific noncovalent complexes and complexes of biomolecules
with metal ions. These species are stable under physiological conditions, but may
not survive sample crystallization or laser desorption and ionization processes.
Up to now, the ability of MALDI to detect specific noncovalent complexes is just
starting to be explored (Cohen et al. 1997; Glocker et al. 1996; Gruic-Sovulj et al.
1997; Woods et al. 1995) and only little work has been done with MALDI (Lehmann et al. 1997; Nelson and Hutchens 1992) and also ESI (Loo 1997; Veenstra et
al. 1998) to compare solution and gas-phase chemistries of noncovalent and
metal ion-biomolecule complexes.
Our aim was therefore to detect the specific complexes of a zinc finger peptide
with Znz+, as well as with Znz+ and oligodeoxynucleotides with MALDI MS. The
challenge in the second case was to transfer relatively weakly bound species
intact into the gas phase. An important aim was also to distinguish between specific and nonspecific aggregation; the latter may take place in the MALDI plume
1 Laboratorium fiir Organische Chemie, ETH Zentrum, Universitatstr. 16, CH-8092 Ziirich,
Switzerland.
Detection of Specific Zinc Finger Peptide Complexes
with Matrix-Assisted Laser Desorption/Ionization Mass
Spectrometry
E. LEHMANN l and R. ZENOBI l
1
Introduction
Noncovalent complexes play an important role in biochemistry and molecular
biology. Various methods have been used to investigate these complexes. Among
them, size exclusion chromatography and gel electrophoresis, for example, are
used for determining stoichiometry and molecular weights. These conventional
methods are relatively time- and material-consuming. A recent analytical tool to
overcome these problems is mass spectrometry (MS), characterized by high sensitivity and speed. Together with electrospray ionization (ESI) mass spectrometry, matrix-assisted laser desorption/ionization (MALDI) mass spectrometry is a
soft ionization technique, i.e. leads to little fragmentation of the analyte. For this
reason, it is a powerful method for the analysis of various biomolecules and synthetic polymers (Bahr et al. 1992; Hillenkamp et al. 1991). MALDI spectra are
generated by laser irradiation of analyte molecules embedded in an excess of a
crystalline matrix. This matrix is responsible for the absorption of the incident
laser light and also plays an important role in analyte ionization.
A relevant question to ask is whether a correspondence exists between
solution-phase behavior and the observed gas-phase ions and how soft MALDI is
for the analysis of specific noncovalent complexes and complexes of biomolecules
with metal ions. These species are stable under physiological conditions, but may
not survive sample crystallization or laser desorption and ionization processes.
Up to now, the ability of MALDI to detect specific noncovalent complexes is just
starting to be explored (Cohen et al. 1997; Glocker et al. 1996; Gruic-Sovulj et al.
1997; Woods et al. 1995) and only little work has been done with MALDI (Lehmann et al. 1997; Nelson and Hutchens 1992) and also ESI (Loo 1997; Veenstra et
al. 1998) to compare solution and gas-phase chemistries of noncovalent and
metal ion-biomolecule complexes.
Our aim was therefore to detect the specific complexes of a zinc finger peptide
with Znz+, as well as with Znz+ and oligodeoxynucleotides with MALDI MS. The
challenge in the second case was to transfer relatively weakly bound species
intact into the gas phase. An important aim was also to distinguish between specific and nonspecific aggregation; the latter may take place in the MALDI plume
1 Laboratorium fiir Organische Chemie, ETH Zentrum, Universitatstr. 16, CH-8092 Ziirich,
Switzerland.