1. Peptide analogues of a subdominant epitope expressed in ebv-associated tumors: synthesis and immunological activity
M Marastoni, M Bazzaro, F Micheletti, R Gavioli, R Tomatis J Med Chem. 2001 Jul 5;44(14):2370-3. doi: 10.1021/jm001136a.
H-Cys-Leu-Gly-Gly-Leu-Leu-Thr-Met-Val-OH (CLG) peptide is an EBV subdominant epitope that represents the target of HLA-A2 restricted CTL responses. The CLG peptide has low affinity for HLA-A2 and does not produce stable complexes, both factors that determine weak CTL responses. In contrast, the [Tyr(1), Ala(3)]CLG (YLA) analogue showed high affinity for HLA-A2 molecules and efficiently stimulated CLG-specific CTL precursors. Nevertheless, this modified epitope showed low enzymatic stability. To further improve the immunotherapeutical potential of this "improved epitope", we have synthesized and tested YLA analogues containing different modifications next to the scissile peptide bond. Among the analogues we found three peptides, with higher enzymatic resistance, that efficiently stimulate CTL responses. These peptides may be used for EBV-specific immunotherapies.
2. TolC of Escherichia coli functions as an outer membrane channel
R Benz, E Maier, I Gentschev Zentralbl Bakteriol. 1993 Apr;278(2-3):187-96. doi: 10.1016/s0934-8840(11)80836-4.
Reconstitution experiments were performed with TolC from Escherichia coli outer membrane by using the lipid bilayer membrane technique. TolC was purified by elution of the oligomeric and the monomeric forms out of preparative SDS-PAGE. The oligomeric but not the monomeric form of the protein was able to increase the specific conductance of artificial lipid bilayer membranes. Investigation of the membrane activity in single-channel experiments suggested that TolC formed ion-permeable channels. The channels of 80 pS in 1 M KCl had a much smaller single-channel conductance than the general diffusion pores of E. coli outer membrane (1500 pS). The single-channel conductance was only moderately dependent on the bulk aqueous KCl concentration which indicated either ion binding or charge effects. Titration of TolC-induced membrane conductance with peptides lead to a dose-dependent decrease of the conductance. This result suggested that TolC contained a binding site for peptides. A dissociation constant of 20 mM was calculated for the binding of the tripeptide H-Gly-Gly-Leu-OH to the binding site. The results are consistent with the assumption that TolC acts as an outer membrane channel for peptides.
3. Energy-resolved mass spectrometry: a comparison of quadrupole cell and cone-voltage collision-induced dissociation
A G Harrison Rapid Commun Mass Spectrom. 1999;13(16):1663-70. doi: 10.1002/(SICI)1097-0231(19990830)13:163.0.CO;2-T.
Collision-induced dissociation (CID) can be effected in the interface region between atmospheric pressure ionization sources and single quadrupole mass analyzers. By varying the electric field in these cone-voltage CID experiments energy-resolved mass spectra can be obtained leading to breakdown graphs exploring the energy evolution of the fragmentation pathways. The breakdown graphs obtained from these cone-voltage CID studies are very comparable to those obtained by varying the collision energy in the quadrupole collision cell of a BEqQ mass spectrometer. The comparison has been made for the protonated peptides H-Leu-Gly-Gly-OH, H-Gly-Leu-Gly-OH, H-Gly-Gly-Leu-OH and Leu-enkephalin
