A number of recent developments in DNA analysis by capillary electrophoresis are here reviewed. They include capillary arrays for fast, parallel DNA sequencing as well as microfabricated capillary arrays. Microfluidic chips for DNA sizing and quantitation are also covered, as well as microdevices containing arrays of regular obstacles acting as size-separators during DNA migration. Screening of DNA point mutations by two much improved techniques is also reported: in one case, such mutations are detected (but only on relative short, ca. 60-70 base-long fragments) by free electrophoresis in rather acidic (pH ca. 3) buffers; in the case of single-strand chain polymorphism, an improved technique is described based on near-neutral pH buffers with mixtures of Tris/MES cations/zwitterions. When studying the behavior of inorganic and organic cations in the Debye-Hückel layer of DNA, it was found that the latter (especially a large number of Good's buffers and other zwitterions, such as His) would bind to the DNA filament not only via charge interaction, but also via additional bonds, notably hydrogen bonds, thus altering the electrophoretic (and possibly the biological) behavior of DNA molecules. However, whether or not borate ions would bind to DNA remains still an unsettled question. Finally, capillary electrophoresis was found to be instrumental in measuring fine physicochemical parameters pertaining to DNA polyelectrolytes, such as their free mobility and their translational diffusion coefficients.

Retinoids are quite insoluble and chemically unstable compounds in thc aqucous mcdium, such that natural retinoids nccd to be bound to specific retinoid-binding protcins to be protected, solubilized and transported in body fluids. All-tmns retilioic acid exhibits a relatively high affinity for thyroxine-binding transthyretin irz vitm and this protein is a good candidate for the transport of retinoic acid adn~iniskred us pharmacological or antitumor agent. To dcfitic structural features esseniial for the recognition by transthyrctin of a ligand which is structurally unrclatcd to thyroxine, we have cocrysiallized humun transthyrctin with retinoic acid and deleiinincd its structure at 0.18-nrri tesulution. The retinoid fits into thc two chemically identical thyroxine-binding sites, which are located in the ccntral channcl that rutis through the tetranieric transthyretin. The cyclohexene ring of the hound retinoid is innermost, occupying the sarrie position of the phenolic ring of the bound 3,3'-diiorlo-L-tliyrotiitie, whereas the carboxylate group, like the same group of the thyroid hormone, participatcs in an ionic interaction with the LyslS sidc chain at the entrancc of the channel. Despitc the fact that transthyretin was cocrystallized with all-/rrrn,r-retinoic acid, the isoprene chain of the bound retinoid hus bccn found i n a non-exlendcd conforniation. This feature, that allows thc carboxylate to oricnt in a manner suitable for ion-pair association with the LyslS side chain, is attributable to the conversion of all-trans-retinoic acid into cis-isomers or folded cotifortiiers. It is concluded that the prcsence, i n an essentially hydrophobic rnolecular corc of the appropriate s i 7~ 01 a negatively charged group at the correct position is a crucial requirement for Iigand-tratisthyretirl recognition. Whereas the binding of the ligand has no remarkable conscqucnces for the prolein structure, alltmns-retinoic acid undergoes structural changes such as to interact favorably with residues preseni in the thyroxine-binding sites, rcsetiibling roughly the natural ligantl.

In the present paper, the covalent immobilisation of the digesting enzyme trypsin has been achieved through photo-immobilisation on a portion of a silica capillary, thus leading to the construction of a capillary electrophoretic (CE)-microreactor for peptide mapping. The CE-microreactor is characterised by being a single piece, thus ensuring no fluidic or electrical leakage. The enzyme was immobilised with a surface density of 15.8 microg/cm(2), the stability was high (80% after 38 days) and the rate of conversion was 0.2 ng/s. On-line protein mapping was tested with proteins of different dimensions, showing competitiveness in terms of time (completed map within 15 min) and exhaustive maps of small proteins. The results of the CE-microreactor and the potential to immobilise biocomponents easily on a desired portion of the capillary indicate further developments towards the construction of a variety of miniaturised enzymatic screening devices for high-throughput screening analysis.

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