Plucking, pillaging and plundering proteomes with combinatorial peptide ligand libraries.
Recent developments in the technique of combinatorial peptide ligand libraries, for enhancing the visibility of the low-abundance proteome, are reviewed here. Novel en bloc elution systems, allowing essentially complete proteome recovery in a single step, are reported here, particularly, en bloc elution with 3-5% boiling sodium dodecyl sulphate (SDS) or in urea-thiourea-CHAPS added with either 40 mM formic acid or 25 mM cysteic acid. Novel capturing systems are also discussed: in particular, although capturing at pH 7.2 in physiological saline has always been recommended, it is shown that capturing also at acidic (pH 3.8) and alkaline (pH 9.5) values substantially increments the total captured protein population. Some examples of detection of novel proteins by the described methodology are also discussed. In particular, in the case of venom proteins, where essentially all components had been detected and fully described by conventional means, the application of the ligand library technology allowed the discovery of two, previously unreported, trace enzymes necessary for the maintenance of the native structure of venom components, namely peroxiredoxin and glutaminyl cyclase. In the case of the red blood cell (RBC) cytoplasmic proteome, where a grand total of 1570 components of the 2% minority proteomes have been identified, these findings allowed to unravel the genetic defect of a rare RBC disease, called congenital dyserythropoietic anemia type II. The mutations are located in the SEC23B gene coding for the SEC23B protein, detected for the first time in the RBC proteome thanks to the peptide capturing technology.
Righetti
Pier Giorgio
PG
Department of Chemistry, Materials and Chemical Engineering Giulio Natta, Politecnico di Milano, Via Mancinelli 7, Milano 20131, Italy. piergiorgio.righetti@polimi.it
Boschetti
Egisto
E
Zanella
Alberto
A
Fasoli
Elisa
E
Citterio
Attilio
A
D000818:Animals; D020650:Combinatorial Chemistry Techniques / Q000379:methods*; D003593:Cytoplasm / Q000737:chemistry; D015180:Electrophoresis, Gel, Two-Dimensional; D004912:Erythrocytes; D006801:Humans; D006863:Hydrogen-Ion Concentration; D019151:Peptide Library; D040901:Proteomics / Q000379:methods*; D012910:Snake Venoms / Q000737:chemistry
D019151:Peptide Library; D012910:Snake Venoms