A dimer between monomers and hexamers-Oligomeric variations in glucosamine-6-phosphate deaminase family.

In bacteria that live in hosts whose terminal sugar is a sialic acid, Glucosamine-6-phosphate deaminase (NagB) catalyzes the last step in converting sialic acid into Fructose-6-phosphate. These bacteria then use the Fructose-6-phosphate as an energy source. The enzyme NagB exists as a hexamer in Gram-negative bacteria and is allosterically regulated.

Automation aided optimization of cloning, expression and purification of enzymes of the bacterial sialic acid catabolic and sialylation pathways enzymes for structural studies.

The process of obtaining a well-expressing, soluble and correctly folded constructs can be made easier and quicker by automating the optimization of cloning, expression and purification. While there are many semiautomated pipelines available for cloning, expression and purification, there is hardly any pipeline that involves complete automation. Here, we achieve complete automation of all the steps involved in cloning and in vivo expression screening.

Carbohydrate polymers for nonviral nucleic acid delivery.

Carbohydrates have been investigated and developed as delivery vehicles for shuttling nucleic acids into cells. In this review, we present the state of the art in carbohydrate-based polymeric vehicles for nucleic acid delivery, with the focus on the recent successes in preclinical models, both in vitro and in vivo. Polymeric scaffolds based on the natural polysaccharides chitosan, hyaluronan, pullulan, dextran, and schizophyllan each have unique properties and potential for modification, and these results are discussed with the focus on facile synthetic routes and favorable performance in biological systems.

Versatile supramolecular pDNA vehicles via "click polymerization" of beta-cyclodextrin with oligoethyleneamines.

Herein, we report the efficient synthesis of high molecular weight polymers (up to 331 kDa) that contain beta-cyclodextrin within the polymer backbone and the examination of these structures for pDNA delivery within cultured mammalian cells. Two series of polymers were synthesized, one with variation in oligoethyleneamine stoichiometry, Cd1(46), Cd2(44), Cd3(49), and Cd4(47) (1-4 oligoethyleneamines in the repeat unit, respectively and similar degree of polymerization, n(w)=44-49) and another with variation in polymer length (four ethyleneamines in the repeat unit), Cd4(27), Cd4(47), Cd4(93), and Cd4(200) [n(w)=27, 47, 93, 200] via the "click reaction". The two series of polymers revealed efficient pDNA binding and compaction through gel electrophoresis, dynamic light scattering, and transmission electron microscopy experiments.

Polycationic beta-cyclodextrin "click clusters": monodisperse and versatile scaffolds for nucleic acid delivery.

Herein, a novel series of multivalent polycationic beta-cyclodextrin "click clusters" with discrete molecular weight have been synthesized, characterized, and examined as therapeutic pDNA carriers. The materials were creatively designed based on a beta-cyclodextrin core to impart a biocompatible multivalent architecture and oligoethyleneamine arms to facilitate pDNA binding, encapsulation, and cellular uptake. An acetylated-per-azido-beta-cyclodextrin (4) was reacted with series of alkyne dendrons (7a-e) (containing one to five ethyleneamine units) using copper-catalyzed 1,3-dipolar cycloaddition, to form a series of click clusters (9a-e) bearing 1,2,3-triazole linkers.

Effects of trehalose click polymer length on pDNA complex stability and delivery efficacy.

Cationic polymers are currently being studied as non-viral vectors to deliver therapeutic DNA into cells. In this study, a series of trehalose-based glycopolymers containing four secondary amines in the repeat unit were synthesized via the 'click reaction' [degrees of polymerization (n(w))=35, 53, 75, or 100] to elucidate how the polymer length affects the bioactivity. The four structures bound and charge-neutralized pDNA with similar affinity that was independent of the length, as determined through gel electrophoresis, heparin competitive displacement, and isothermal titration calorimetric assays.

Trehalose click polymers inhibit nanoparticle aggregation and promote pDNA delivery in serum.

Herein, three new glycopolymers have been synthesized via "click polymerization" to promote nucleic acid delivery in the presence of biological media containing serum. These structures were designed to contain a trehalose moiety to promote biocompatibility, water solubility, and stability against aggregation, amide-triazole groups to enhance DNA binding affinity, and an oligoamine unit to facilitate DNA encapsulation, phosphate neutralization, and interactions with cell surfaces. A 2,3,4,2',3',4'-hexa-O-acetyl-6,6'-diazido-6,6'-dideoxy-D-trehalose (4) monomer was polymerized via copper(I)-catalyzed azide-alkyne cycloaddition with a series of dialkyne-amide comonomers that contain either one, two, or three Boc-protected secondary amines (7a, 7b, or 7c, respectively).