The growth of marine Pseudomonas sp. NCIMB 2021 as continuous cultures in the presence of surfaces of AlSl 316 stainless steel allowed the isolation and partial chemical characterization of exopolymers released into the culture medium (free exopolymers), as well as capsular and biofilm exopolymers. Fourier-transform infrared (FTIR) spectroscopy demonstrated the presence of 0-and N-acetylation within the carbohydrate moieties and a predominant 3,,-helical structure of the protein component, highly resistant to hydrogeddeuterium exchange. Differences between the exopolymers were apparent. Relatively less uronic acid residues were detected in the capsular exopolymers compared to either the biofilm or free exopolymers. 0-and Nacetylation were greatest in the biof ilm exopolymer. SDS-PAGE protein profiles confirmed differences between exopolymers. The secondary structures of proteins determined using FTIR spectroscopy indicated that the capsular exopolymer had reduced helical content and an increased aggregated strand content compared to the biofilm exopolymer. However, the free exopolymer had an increased P-sheet component and a reduced unordered component when compared to the biofilm and capsular exopolymers. These data suggest that exopolymer chemistry varies with cellular mode of growth.
Directed enzyme prodrug therapy is an extensive area of research in cancer chemotherapy. Although very promising, the current directed approaches are still hampered by inefficient enzyme expression and tumor targeting. This work investigates the viability of using metal nanoparticles as a novel delivery vehicle for prodrug-activating enzymes. Using genetically incorporated amino acid sequences, a nitroreductase from E. coli was directly immobilized onto a 50 nm gold colloid, as confirmed by gel electrophoresis, DLS, and UV-vis spectroscopy. The resulting conjugates showed excellent stability in changing proton and sodium chloride environments, including PBS at 37 °C. Remarkably, the immobilized nitroreductase retained more than 99% activity to the CB1954 prodrug without the need for stabilizers. This work provides the foundation for attaching prodrug-activating enzymes to metal nanoparticles for future use in directed enzyme prodrug therapy.
Organised multilayers were formed from the controlled self-assembly of ferrocene alkyl thiols on Au(111) surfaces. The control was accomplished by increasing the concentration of the thiol solutions used for the assembly. Cyclic voltammetry, ellipsometry, scanning probe microscopy (STM and AFM) and in situ FTIR spectroscopy were used to probe the di †erences between mono-and multilayers of the same compounds. Electrochemical desorption studies conÐrmed that the multilayer structure is attached to the surface via one monolayer. The electrochemical behaviour of the multilayers indicated the presence of more than one controlling factor during the oxidation step, whereas the reduction was kinetically controlled which contrasts with the behaviour of monolayers, which exhibit kinetic control for the oxidation and reduction steps. Conventional and imaging ellipsometry conÐrmed that multilayers with well-deÐned increments in thickness could be produced. However, STM indicated that at the monolayer stage, the thiols used promote the mobility of Au atoms on the surface. It is very likely that the multilayer structure is held together through hydrogen bonding. To the best of out knowledge, this is the Ðrst example of a controlled one-step growth of multilayers of ferrocenyl alkyl thiols using self-assembly techniques.
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