Synthesis and application of cleavable and hydrophilic crosslinking reagents

Abstract

Bifunctional imidoesters are due to their mild reaction especially suitable for the crosslinking of proteins. Most often used are diimidates with a medium span (4 or 6 CH2-groups). Reagents with a wider span might be of interest. In such a case, however, the bridge should be more hydrophilic. Bifunctional imidoesters have, therefore, been prepared from dinitriles NC-(CH2)2-X-(CH2)2-CN; X being -O-, -O-(CH2)n-O- or -O-(CH2)m-O-(CH2)n-O-. The bridge of diimidoesters can also be labelled by coloured or fluorescent groups. Diimidoesters containing -S-S-bonds in the bridge can be cleaved more easily after their reaction with proteins. Dimethyl-3.3' -(8- delta -dithiahexamethylenedioxy)-dipropionimidate, a "long", cleavable and more hydrophilic diimidate, is a promising new reagent. In cases, where HS-(CH2)3-C(=NH)-OCH3- HC1 is is used for crosslinking of proteins, this reagent can possibly be replaced by 2-iminothiolane hydrochloride, a cyclic thioimidate, which is easier to prepare and to handle.