In this study, structure changes of regenerated cellulose fibers wet-spun from a cotton linter pulp (degree of polymerization approximately 620) solution in an NaOH/urea solvent under different conditions were investigated by simultaneous synchrotron wide-angle X-ray diffraction (WAXD) and small-angle X-ray scattering (SAXS). WAXD results indicated that the increase in flow rate during spinning produced a better crystal orientation and a higher degree of crystallinity, whereas a 2-fold increase in draw ratio only affected the crystal orientation. When coagulated in a H2SO4/Na2SO4 aqueous solution at 15 degrees C, the regenerated fibers exhibited the highest crystallinity and a crystal orientation comparable to that of commercial rayon fibers by the viscose method. SAXS patterns exhibited a pair of meridional maxima in all regenerated cellulose fibers, indicating the existence of a lamellar structure. A fibrillar superstructure was observed only at higher flow rates (>20 m/min). The conformation of cellulose molecules in NaOH/urea aqueous solution was also investigated by static and dynamic light scattering. It was found that cellulose chains formed aggregates with a radius of gyration, Rg, of about 232 nm and an apparent hydrodynamic radius, Rh, of about 172 nm. The NaOH/urea solvent system is low-cost and environmentally friendly, which may offer an alternative route to replace more hazardous existing methods for the production of regenerated cellulose fibers.

Reverse transcription in hepadnaviruses is primed by the viral reverse transcriptase (RT) (protein priming) and requires the interaction between the RT and a specific viral RNA template termed . Protein priming is resistant to a number of RT inhibitors that can block subsequent viral DNA elongation and likely requires a distinct "priming" conformation. Furthermore, protein priming may consist of two distinct stages, i.e., the attachment of the first deoxynucleotide to RT (initiation) and the subsequent addition of 2 or 3 deoxynucleotides (polymerization). In particular, a truncated duck hepatitis B virus RT (MiniRT2) Hepadnaviruses (hepatitis B viruses [HBVs]) are retroid viruses that replicate a small (ca. 3-kb) DNA genome via an RNA intermediate, the pregenomic RNA (pgRNA). The hepadnavirus reverse transcription pathway shows both similarities to and differences from that of classical retroviruses, including the mechanisms of nucleocapsid assembly and the initiation of DNA synthesis (40,42,45). To carry out their unique life cycle, the hepadnaviruses encode a specialized reverse transcriptase (RT) protein that displays a number of unique properties distinct from those of its retrovirus counterparts (23). Chief among these is its ability to initiate DNA synthesis de novo without the help of any DNA or RNA primer. Instead, a specific tyrosine residue within RT itself is used as the primer to initiate viral minus strand DNA synthesis (the so-called protein priming reaction) (31,32,52,53,56,58).The unique ability of the hepadnavirus RT to carry out protein priming by using itself as a protein primer is reflected in its novel structural organization (11,22,23,37). RT consists of four domains: from the N to the C terminus, they are the terminal protein (TP), the spacer, the RT, and the RNase H domains. TP is conserved among all hepadnaviruses but absent from any other known proteins. It is within TP where the aforementioned primer tyrosine residue is located. The functionally dispensable spacer connects TP to the central RT domain. Both the RT and RNase H domains share sequence homology with conventional RTs, including the YMDD active site motif in the RT domain and the catalytic D residues in the RNase H domain. Based on sequence alignment, structure modeling, and some genetic and biochemical data, the RT domain can be further divided into the finger, palm, and thumb subdomains, as in virtually all DNA and RNA polymerases (14,41).Although no viral protein other than RT itself is required for protein priming in hepadnaviruses, there is an absolute requirement for a specific RNA template, the short RNA stemloop structure, called ε, located on pgRNA (36, 53). The ε RNA bears two inverted repeat sequences and can fold into a conserved stem-loop structure, with a lower and an upper stem, an apical loop, and an internal bulge (16,17,22,25,28). Using the internal bulge of ε as the obligatory template and the specific tyrosine residue within its TP domain as the protein primer, RT synthesizes a 3-to 4-nucleotide (nt)-long...

Oxidative stresses triggered by reactive oxygen species (ROS) that damage various cellular components are unavoidable for virtually all living organisms. In defense, microorganisms have evolved sophisticated mechanisms to sense, respond to, and battle against ROS. Shewanella oneidensis, an important research model for applied and environmental microbes, employs OxyR to mediate the response to H 2 O 2 by derepressing the production of the major H 2 O 2 scavenger KatB as a major means toward these goals. Surprisingly, despite enhanced H 2 O 2 degradation, the oxyR mutant carries a viability deficiency phenotype (plating defect), which can be suppressed by the addition of exogenous iron species. Experiments showed that the defect was not due to iron starvation. Rather, multiple lines of evidence suggested that H 2 O 2 generated abiotically in lysogeny broth (LB) is responsible for the defect by quickly killing mutant cells. We then showed that the iron species suppressed the plating defect by two distinct mechanisms, either as an H 2 O 2 scavenger without involving living cells or as an environmental cue to stimulate an OxyR-independent response to help cells cope with oxidative stress. Based on the suppression of the plating defect by overproduction of H 2 O 2 scavengers in vivo, we propose that cellular components that are vulnerable to H 2 O 2 and responsible for the defect may reside outside the cytoplasm. IMPORTANCEIn bacteria, OxyR is the major regulator controlling the cellular response to H 2 O 2 . The loss of OxyR results in reduced viability in many species, but the underlying mechanism is unknown. We showed in S. oneidensis that this defect was due to H 2 O 2 generated abiotically in LB. We then showed that this defect could be corrected by the addition of Fe 2؉ or catalase to the LB or increased intracellular production of catalase. Further analyses revealed that Fe 2؉ was able not only to decompose H 2 O 2 directly but also to stimulate the activity of OxyR-independent H 2 O 2 -scavenging enzymes. Our data indicate that iron species play a previously underappreciated role in protecting cells from H 2 O 2 in environments.

SummaryShewanella oneidensis is renowned for its respiratory versatility, which is largely due to abundant c-type cytochromes. Maturation of these proteins depends on a Ccm system encoded by genes in an unusual chromosomal arrangement, but the detailed mechanism is not understood. In this study, we identify SO0265 as CcmI, an apocytochrome c chaperone that is important and essential for maturation of c-type cytochromes with the canonical heme binding motif(s) (HBM; CX2CH) and nitrite reductase NrfA carrying a non-canonical CX2CK motif respectively. We show that the N-terminal transmembrane segment of CcmI, CcmI-1, is sufficient for maturation of the former but the entire protein is required for maturation of the latter. Although S. oneidensis possesses a heme lyase, SirEFG, dedicated for non-canonical HBMs, it is specific for SirA, a sulfite reductase with a CX15CH motif. By presenting evidence that the periplasmic portion of CcmI, CcmI-2, interacts with NrfA, we suggest that CcmI also takes the role of Escherichia coli NrfG for chaperoning apo-NrfA for maturation at CX2CK. Moreover, intact CcmI is required for maturation of NrfA, presumably by ensuring that heme attachment at canonical HBMs occurs before apoprotein degradation.

No abstract

Antibiotic resistance is a significant crisis that threatens human health and safety worldwide. There is an urgent need for new strategies to control multidrug-resistant (MDR) bacterial infections. The latest breakthrough in gene-editing tools based on CRISPR/Cas9 has potential application in combating MDR bacterial infections because of their high targeting ability to specifically disrupt the drug resistance genes that microbes use for infection or to kill the pathogen directly. Despite the potential that CRISPR/Cas9 showed, its further utilization has been hampered by undesirable delivery efficiency in vivo. Nanotechnology offers an alternative way to overcome the shortcomings of traditional delivery methods of therapeutic agents. Advances in nanotechnology can improve the efficacy and safety of CRISPR/Cas9 components by using customized nanoparticle delivery systems. The combination of CRISPR/Cas9 and nanotechnology has the potential to open new avenues in the therapy of MDR bacterial infections. This review describes the recent advances related to CRISPR/Cas9 and nanoparticles for antimicrobial therapy and gene delivery, including the improvement in the packaging and localizing efficiency of the CRISPR/Cas9 components in the NP (nanoparticle)/CRISPR system. We pay particular attention to the strengths and limitations of the nanotechnology-based CRISPR/Cas9 delivery system to fight nosocomial pathogens.We highlight the need for more scientific research to explore the combinatorial efficacy of various nanoparticles and CRISPR technology to control and prevent antimicrobial resistance.

It is well established that OxyR functions as a transcriptional activator of the peroxide stress response in bacteria, primarily based on studies on Escherichia coli.

A reversible thermo-responsive gel system, consisting of Pluronic copolymer mixture of F87 and F127, has been used to successfully carry out the separation of oligonucleotides, for the first time, by microchip-based capillary electrophoresis. Pluronic triblock copolymers F87 (E(61)P(40)E(61)) and F127 (E(99)P(69)E(99)), with E, P, and subscript denoting oxyethylene, oxypropylene, and segment length respectively, have a unique temperature dependent viscosity-adjustable property and a dynamic coating ability in aqueous solution, including 1 x TBE buffer. The mixture solution has a reversible thermo-responsive property and its sol-gel transition temperature can be adjusted ranging from about 17 degrees C to 38 degrees C by varying the relative weight ratio of F87 and F127 at an optimized concentration of approximately 30% (w/v) for oligonucleotide separations. Oligonucleotide sizing markers ranging from 8 to 32 base could be successfully separated in a 1.5 cm long separation channel by the mixture solution in its gel-like state. A 30% (w/v) with a F87/F127 weight ratio of 1 ratio 2 which has a "sol-gel" transition point of about 26 degrees C shows the best sieving ability. The sieving ability of the mixture solution was further confirmed in an Agilent Bioanalyzer 2100 system. Fast separation of oligonucleotides has been achieved within 40 s with one base resolution.