Publikace detail

Chip-based bioaffinity reactor with immobilized anhydrotrypsin for isolation of specific tryptic peptides of human neurotensin

Autoři: Korecká Lucie | Křenková Jana | Minc Nicolas | Viovy jean Louis | Bílková Zuzana

Rok: 2005

Druh publikace: článek ve sborníku

Název zdroje: Mezinárodní sjezd chemické společnosti

Název nakladatele: Slovenská Chemická spoločnost

Místo vydání: Bratislava, Slovensko

Strana od-do: 267-268

Tituly:

Jazyk	Název	Abstrakt	Klíčová slova
cze	Bioafinitní reaktor založený na imobilizovaném anhydrotrypsinu, izolace neurotensinových peptidů	Bioafinitní reaktor založený na imobilizovaném anhydrotrypsinu, izolace neurotensinových peptidů.	bioafinitni reaktor, micro-chip, izolace, peptidy, neurotensin
eng	Chip-based bioaffinity reactor with immobilized anhydrotrypsin for isolation of specific tryptic peptides of human neurotensin	One of the modern trends in biochemistry is focused on the utilization of microfluidic device for protein analysis due to many advantages (low sample consumption, highthroughput analysis etc.). Functional spherical nanoparticles with controlled tendency to aggregate and controlled level of nonspecific sorption could be focused in channel of magnetically active microfluidic device for isolation or enzyme modification of target protein directly in a channel of microfluidic device. Biospecific carrier prepared by immobilization of anhydrotrypsin on magnetic nanoparticles was used for purification, isolation and preconcentration of specific peptides from complex mixture of tryptic peptides in one step. IMER (immobilized magnetic enzyme reactor) with trypsin in tandem with chip-based IMAR (immobilized magnetic affinity reactor) with anhydrotrypsin was used for simplification of peptide mapping including the digestion of protein coupling with purification and preconcentration of peptides for following analysis by HPLC and MS. Anhydrotrypsin is catalytically inactive derivative of bovine trypsin in which serine residue (Ser-195) is first modified with phenylmethanesulfonyl fluoride (PMSF), an active site-directed reagent, and then the modified protein is treated with alkali results. Anhydrotrypsin enables highly specific trapping of peptides which correspond to the products generated by the action of various trypsin-like proteases and binds, in slightly acidic conditions, only specific peptides containing arginine and lysine residues at their C-termini2. High purity and preconcentration of peptides isolated on immobilized anhydrotrypsin significantly facilitated the identification of protein fingerprint even for protein in submicromolar concentration.	bioaffinity reactor; microchip; isolation; peptides; neurotensin

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