Peptide and protein separations by capillary electrophoresis in the presence of mono- and diquaternarized diamines.
Two compounds, derivatives of 1,4-diazobicyclo[2,2,2]octane (DABCO), have been evaluated as potential quenchers of silanol interactions with peptides and proteins during their capillary zone electrophoresis (CZE) separations. They are: 1-(4-iodobutyl)4-aza-1-azoniabicyclo[2,2,2]octane iodide (M7C4I) and 1,4-didecyl-1,4-diazoniabicyclo [2,2,2]octane dibromide (C10M7C10). The first compound is known to react with the wall, by forming a covalent bond via alkylation of silanols. On the contrary, the second one (C10M7C10) can only loosely interact with silica due to lack of reactive iodine and to a much too short distance (a C(2)) between the two quaternary nitrogens. Very good peptide maps of protein digests can be obtained in isoelectric glutamic acid (Glu) buffer, at pH 3.52 by utilizing the M7C4I. However, in the case of total tissue extracts, excellent resolution is obtained only with the first eluting part of the analyte spectrum (i.e., peptides and smaller proteins). With larger proteins, interaction with the wall and loss of resolution is experienced. When using the C10M7C10, good resolution of di- and tripeptides is obtained, while a loss of resolution is observed with entire protein digest. M7C4I does not seem to interact with the peptide/protein analytes, and simply repels them from the wall via its positive charges; it is believed that disalt (C10M7C10) acts by interacting with the same compounds, possibly by forming micelles in solution.
Pontoglio
Alessandro
A
Institute of Molecular Bioimaging and Physiology, CNR, Segrate, Milano, Italy.
Viganò
Agnese
A
Sebastiano
Roberto
R
Citterio
Attilio
A
Maragnoli
Luca
L
Righetti
Pier Giorgio
PG
Gelfi
Cecilia
C
D000818:Animals; D003959:Diamines / Q000032:analysis*; D019075:Electrophoresis, Capillary / Q000379:methods*; D010449:Peptide Mapping; D010455:Peptides / Q000302:isolation & purification*; D011506:Proteins / Q000302:isolation & purification*; D051381:Rats; D017207:Rats, Sprague-Dawley
D003959:Diamines; D010455:Peptides; D011506:Proteins