Sprayed water-based lignin colloidal nanoparticle-cellulose nanofibril hybrid films with UV-blocking ability.

In the context of global climate change, the demand for new functional materials that are sustainable and environmentally friendly is rapidly increasing. Cellulose and lignin are the two most abundant raw materials in nature, and are ideal components for functional materials. The hydrophilic interface and easy film-forming properties of cellulose nanofibrils make them excellent candidates for natural biopolymer templates and network structures. Lignin is a natural UV-shielding material, as it contains a large number of phenolic groups. In this work, we have applied two routes for spray deposition of hybrid films with different laminar structures using surface-charged cellulose nanofibrils and water-based colloidal lignin particles. As the first route, we prepare stacked colloidal lignin particles and cellulose nanofibrils hybrid film through a layer-by-layer deposition. As the second route, we spray-deposite premixed colloidal lignin particles and cellulose nanofibrils dispersion to prepare a mixed hybrid film. We find that cellulose nanofibrils act as a directing agent to dominate the arrangement of the colloidal lignin particles in a mixed system. Additionally, cellulose nanofibrils eliminate the agglomerations and thus increase the visible light transparency while retaining the UV shielding ability. Our research on these colloidal lignin and cellulose nanofibril hybrid films provides a fundamental understanding of using colloidal lignin nanoparticles as functional material on porous cellulose-based materials, for example on fabrics.

(S,2S,3R)-2-(2-Methyl-propane-2-sulfin-amido)-3-phenyl-butyronitrile.

The absolute configuration has been determined for the title compound, C(14)H(20)N(2)OS. Inter-molecular N-H⋯O hydrogen bonds are observed in the crystal packing, forming infinitive one-dimensional chains with the base vector [100].

(S,2S,3S)-2-(2-Methyl-propan-2-sulfin-amido)-3-phenyl-butyronitrile.

The absolute configuration has been determined for the title compound, C(14)H(20)N(2)OS. There are two independent mol-ecules in the asymmetric unit. Inter-molecular N-H⋯O hydrogen bonds are observed in the crystal packing, forming infinite chains with the base vectors [100] and [010]. Each chain contains only one of the two independent mol-ecules.

Tuning the Density of Zwitterionic Polymer Brushes on PET Fabrics by Aminolysis: Effect on Antifouling Performances.

Here, we synthesize zwitterionic polymer brushes on polyester fabrics by atom transfer radical polymerization (ATRP) after a prefunctionalization step involving an aminolysis reaction with ethylenediamine. Aminolysis is an easy method to achieve homogeneous distributions of functional groups on polyester fibers (PET) fabrics. Varying the polymerization time and the prefunctionalization conditions of the reaction, it is possible to tune the amount of water retained over the surface and study its effect on protein adhesion. This study revealed that the polymerization time plays a major role in preventing protein adhesion on the PET surface.

Nonfouling textiles with tunable antimicrobial activity based on a zwitterionic polyamine finish.

Antimicrobial finishes for textiles and other surfaces that act without the release of biocides to the environment (contact biocides) or by inhibiting microbial adhesion (antifouling action) are viewed as promising and environmentally friendly alternatives to current products. We have used polyvinylamine polymers that were functionalized with zwitterionic sulfobetaine side chains with different degrees of substitution (DS) for the finishing of poly(ethylene terephthalate) (PET) and cotton fabrics in a water-based pad-dry-cure process. After washing with different surfactants, a stable finish with total polymer add-ons of 0.2-0.5 wt% was achieved. The finished textiles efficiently inhibited the adhesion of proteins and bacteria to the surface even with a small DS as low as 20%. Textiles finished with polymers with a low DS also showed significant antibacterial activity, most notably against Staphylococcus aureus. Accordingly, textile finishes with either pure antiadhesive (DS > 50%) or combined antiadhesive and antibacterial properties (DS = 20-50%) are accessible using this approach.

Harnessing the chemical activation inherent to carrier protein-bound thioesters for the characterization of lipopeptide fatty acid tailoring enzymes.

Here, we report a new experimental approach utilizing an amide ligation reaction for the characterization of acyl carrier protein (ACP)-bound reaction intermediates, which are otherwise difficult to analyze by traditional biochemical methods. To explore fatty acid tailoring enzymes of the calcium-dependent antibiotic (CDA) biosynthetic pathway, this strategy enabled the transformation of modified fatty acids, covalently bound as thioesters to an ACP, into amide ligation products that can be directly analyzed and compared to synthetic standards by HPLC-MS. The driving force of the amide formation is the thermodynamic activation inherent to thioester-bound compounds. Using this novel method, we were able to characterize the ACP-mediated biosynthesis of the unique 2,3-epoxyhexanoyl moiety of CDA, revealing a new type of FAD-dependent oxidase HxcO with intrinsic enoyl-ACP epoxidase activity, as well as a second enoyl-ACP epoxidase, HcmO. In general, our approach should be widely applicable for the in vitro characterization of other biosynthetic systems acting on carrier proteins, such as integrated enzymes from NRPS and PKS assembly lines or tailoring enzymes of fatty and amino acid precursor synthesis.

AknT is an activating protein for the glycosyltransferase AknS in L-aminodeoxysugar transfer to the aglycone of aclacinomycin A.

During biosynthesis of the anthracycline antitumor agents daunomycin, adriamycin, and aclacinomycin, the polyketide-derived tetracyclic aglycone is enzymatically glycosylated at the C7-OH by dedicated glycosyltransferases (Gtfs) that transfer L-2,3,6-trideoxy-3-aminohexoses. In aclacinomycins, the first deoxyhexose is predicted to be transferred via AknS action, then subjected to further elongation to a trisaccharide by the subsequent Gtf, AknK. We report here that purified AknS has very low activity in the absence of the adjacently encoded AknT; however, at a 3:1 ratio, AknT stimulates AknS k(cat) by 40-fold up to 0.22 min(-1) for transfer of L-2-deoxyfucose (2-dF) to the aglycone aklavinone. It is likely that several other Gtfs that glycosylate polyketide aglycones also act as two-component catalytic systems. Incubations of purified AknS/AknT/AknK with two aglycones and two dTDP-2-deoxyhexoses produced previously uncharacterized anthracycline disaccharides.

Tailoring of glycopeptide scaffolds by the acyltransferases from the teicoplanin and A-40,926 biosynthetic operons.

The teicoplanin acyltransferase (Atf) responsible for N-acylation of the glucosamine moiety to create the teicoplanin lipoglycopeptide scaffold has recently been identified. Here we use that enzyme (tAtf) and the cognate acyltransferase from the related A-40,926 biosynthetic cluster (aAtf) to evaluate specificity for glycopeptide scaffolds and for the acyl-CoA donor. In addition to acylation of 2-aminoglucosyl glycopeptide scaffolds with k(cat) values of 400-2000 min(-1), both Atfs transfer acyl groups to regioisomeric 6-aminoglucosyl scaffolds and to glucosyl scaffolds at rates of 0.2-0.5 min(-1) to create variant lipoglycopeptides. Using the teicoplanin glycosyltransferase tGtfA, tAtf, and GtfD, a glycosyltransferase from the vancomycin producer, it is possible to assemble a novel lipoglycopeptide with GlcNAc at beta-OH-Tyr(6) and an N(6)-acyl-glucosaminyl-vancosamine at Phegly(4). This study illustrates the utility of chemo- and regioselective acyltransferases and glycosyltransferases to create novel lipoglycopeptides.

Incorporation of glucose analogs by GtfE and GtfD from the vancomycin biosynthetic pathway to generate variant glycopeptides.

Analogs of the glycopeptide antibiotics vancomycin and teicoplanin with alterations in one or both sugar moieties of the disaccharide have been prepared by tandem action of the vancomycin pathway glycosyltransferases GtfE and GtfD. All four regioisomers (2-, 3-, 4-, 6-) of TDP-deoxyglucoses and UDP/TDP-aminoglucoses were prepared, predominantly by action of D-glucopyranosyl-1-phosphate thymidylyltransferase, E(p). GtfE transferred the deoxyglucoses or aminoglucoses onto the 4-OH of 4-hydroxyphenylglycine of both the vancomycin and teicoplanin aglycone scaffolds. Kinetic analysis indicated the 2-, 3-, 4-, and 6-amino-glucoses were transferred by GtfE with only a 4- to 30-fold drop in k(cat) and no effect on K(m) compared to the native substrate, UDP/TDP-glucose, suggesting preparative utility. The next enzyme, GtfD, could utilize the variant glucosyl-peptides as substrates for transfer of L-4-epi-vancosamine. The aminosugar moieties in these variant glycopeptides introduce sites for acylation or reductive alkylation.

A practical method for the stereoselective generation of beta-2-deoxy glycosyl phosphates.

Beta-2-Deoxy sugar nucleotides are substrates used by a variety of glycosyltransferases (Gtfs). We have developed a chemical route to synthesize beta-2-deoxy sugar phosphates that starts from alpha-glycosyl chlorides. Our approach reliably provides access to a range of NDP beta-2-deoxy sugars essential for studying glycosyltransferases involved in the synthesis of biologically active natural products.

Synthesis of linear beta-(1 --> 4)-galacto-hexa- and heptasaccharides and studies directed towards cyclogalactans.

Synthesis of an exclusively beta-(1 --> 4)-linked galactohexa- and heptasaccharide is described by coupling a 2-O-pivaloyl-3,6-O-allyl-protected thiogalactobioside donor with an equally protected, yet terminally 4-OH-free galactopentaoside. The same approach though failed to elaborate cyclic oligomers, as neither cyclodimerization of the correspondingly protected thiogalactotriosides with a 4"-OH could be effected, nor intramolecular glycosidation of the respective hexa- and heptagalactosides with an unprotected 4-OH at one, and phenylthio or sulfoxido groups at the reducing end. The causative factors underlying this are attributed to an inadequate predisposition of the linear beta-(1 --> 4)-galactan chains to adopt the tightly coiled molecular geometry necessary for cyclization--at least at the hexa- and heptasaccharide stage.

Metal--Metal "Communication" of Rh or Pd with Nd in Novel Heterobinuclear Complexes.

The reaction of a neodymium "ate" complex and an electron-rich transition metal chloride by salt elimination is an efficient method for synthesizing heterobinuclear compounds which contain a lanthanide and a Group 9 or 10 metal [Eq. (1), H2 Ap=2-amino-4-methylpyridine]. The use of bisaminopyridinato ligands allows extremely short distances between Rh or Pd and Nd.

The siderophore binding protein FeuA shows limited promiscuity toward exogenous triscatecholates.

Iron acquisition by siderophores is crucial for survival and virulence of many microorganisms. Here, we investigated the binding of the exogenous siderophore ferric enterobactin and the synthetic siderophore mimic ferric mecam by the triscatecholate binding protein FeuA from Bacillus subtilis at the atomic level. The structural complexes provide molecular insights into the capture mechanism of FeuA for exogenous and synthetic siderophores. The protein-ligand complexes show an exclusive acceptance of Λ-stereoconfigured substrates. Ligand-induced cross-bridging of the complexes was not observed, revealing a different thermodynamic behavior especially of the ferric mecam substrate, which was previously shown to dimerize with the enterobactin binding protein CeuE. The nearly identical overall domain movement of FeuA upon binding of ferric enterobactin or ferric mecam compared with endogenously derived ferric bacillibactin implies the importance of the conserved domain rearrangement for recognition by the transmembrane permease FeuBC, for which the conserved FeuA residues E90 and E221 were proved to be essential.

The total synthesis of moenomycin A.

Moenomycin A is the only known natural antibiotic that inhibits bacterial cell wall synthesis by binding to the transglycosylases that catalyze formation of the carbohydrate chains of peptidoglycan. We report here the total synthesis of moenomycin A using the sulfoxide glycosylation method. A newly discovered byproduct of sulfoxide reactions was isolated that resulted in substantial loss of the glycosyl acceptor. A general method to suppress this byproduct was introduced, which enabled the glycosylations to proceed efficiently. The inverse addition protocol for sulfoxide glycosylations also proved essential in constructing some of the glycosidic linkages. The synthetic route is flexible and will allow for derivatives to be constructed to further analyze moenomycin A's mechanism of action.

Characterization of the aminocoumarin ligase SimL from the simocyclinone pathway and tandem incubation with NovM,P,N from the novobiocin pathway.

Simocyclinone D(8) consists of an anguicycline C-glycoside tethered by a tetraene diester linker to an aminocoumarin. Unlike the antibiotics novobiocin, clorobiocin, and coumermycin A(1), the phenolic hydroxyl group of the aminocoumarin in simocyclinone is not glycosylated with a decorated noviosyl moiety that is the pharmacophore for targeting bacterial DNA gyrase. We have expressed the Streptomyces antibioticus simocyclinone ligase SimL, purified it from Escherichia coli, and established its ATP-dependent amide bond forming activity with a variety of polyenoic acids including retinoic acid and fumagillin. We have then used the last three enzymes from the novobiocin pathway, NovM, NovP, and NovN, to convert a SimL product to a novel novobiocin analogue, in which the 3-prenyl-4-hydroxybenzoate of novobiocin is replaced with a tetraenoate moiety, to evaluate antibacterial activity.

A systematic investigation of the synthetic utility of glycopeptide glycosyltransferases.

Glycosyltransferases involved in the biosynthesis of bacterial secondary metabolites may be useful for the generation of sugar-modified analogues of bioactive natural products. Some glycosyltransferases have relaxed substrate specificity, and it has been assumed that promiscuity is a feature of the class. As part of a program to explore the synthetic utility of these enzymes, we have analyzed the substrate selectivity of glycosyltransferases that attach similar 2-deoxy-L-sugars to glycopeptide aglycons of the vancomycin-type, using purified enzymes and chemically synthesized TDP beta-2-deoxy-L-sugar analogues. We show that while some of these glycopeptide glycosyltransferases are promiscuous, others tolerate only minor modifications in the substrates they will handle. For example, the glycosyltransferases GtfC and GtfD, which transfer 4-epi-L-vancosamine and L-vancosamine to C-2 of the glucose unit of vancomycin pseudoaglycon and chloroorienticin B, respectively, show moderately relaxed donor substrate specificities for the glycosylation of their natural aglycons. In contrast, GtfA, a transferase attaching 4-epi-L-vancosamine to a benzylic position, only utilizes donors that are closely related to its natural TDP sugar substrate. Our data also show that the spectrum of donors utilized by a given enzyme can depend on whether the natural acceptor or an analogue is used, and that GtfD is the most versatile enzyme for the synthesis of vancomycin analogues.

Differential inhibition of Staphylococcus aureus PBP2 by glycopeptide antibiotics.

The glycopeptide antibiotics prevent maturation of the bacterial cell wall by binding to the terminal d-alanyl-d-alanine moiety of peptidoglycan precursors, thereby inhibiting the enzymes involved in the final stages of peptidoglycan synthesis. However, there are significant differences in the biological activity of particular glycopeptide derivatives that are not related to their affinity for d-Ala-d-Ala. We compare the ability of vancomycin and a set of clinically relevant glycopeptides to inhibit Staphylococcus aureus PBP2 (penicillin binding protein), the major transglycosylase in a clinically relevant pathogen, S. aureus. We report experiments suggesting that activity differences between glycopeptides against this organism reflect a combination of substrate binding and secondary interactions with key enzymes involved in peptidoglycan synthesis.