Natronoflexus pectinivorans gen. nov. sp. nov., an obligately anaerobic and alkaliphilic fermentative member of Bacteroidetes from soda lakes

Anaerobic enrichment with pectin at pH 10 and moderate salinity inoculated with sediments from soda lakes of the Kulunda Steppe (Altai, Russia) resulted in the isolation of a novel member of the Bacteroidetes, strain AP1T. The cells are long, flexible, Gram-negative rods forming pink carotenoids. The isolate is an obligate anaerobe, fermenting various carbohydrates to acetate and succinate. It can hydrolyze and utilize pectin, xylan, starch, laminarin and pullulan as growth substrates. Growth is possible in a pH range from 8 to 10.5, with an optimum at pH 9.5, and at a salinity range from 0.1 to 2 M Na+. Phylogenetic analysis based on 16S rRNA sequences placed the isolate into the phylum Bacteroidetes as a separate lineage within the family Marinilabilaceae. On the basis of distinct phenotype and phylogeny, the soda lake isolate AP1T is proposed to be assigned in a new genus and species Natronoflexus pectinivorans (=DSM24179T = UNIQEM U807T)

Photodynamic treatment of human breast and prostate cancer cells using rose bengal-encapsulated nanoparticles

Cancer, a prominent cause of death, presents treatment challenges, including high dosage requirements, drug resistance, poor tumour penetration and systemic toxicity in traditional chemotherapy. Photodynamic therapy, using photosensitizers like rose bengal (RB) with a green laser, shows promise against breast cancer cells in vitro. However, the hydrophilic RB struggles to efficiently penetrate the tumour site due to the unique clinical microenvironment, aggregating around rather than entering cancer cells. In this study, we have synthesized and characterized RB-encapsulated chitosan nanoparticles with a peak particle size of ~200 nm. These nanoparticles are readily nternalized by cells and, in combination with a green laser (λ = 532 nm) killed 94–98% of cultured human breast cancer cells (MCF-7) and prostate cancer cells (PC3) at a low dosage (25 μg/mL RB-nanoparticles, fluence ~126 J/cm2, and irradiance ~0.21 W/cm2). Furthermore, these nanoparticles are not toxic to cultured human normal breast cells (MCF10A), which opens an avenue for translational applications

The new role for an old kinase : protein kinase CK2 regulates metal ion transport

The pleiotropic serine/threonine protein kinase CK2 was the first kinase discovered. It is renowned for its role in cell proliferation and anti-apoptosis. The complexity of this kinase is well reflected by the findings of past decades in terms of its heterotetrameric structure, subcellular location, constitutive activity and the extensive catalogue of substrates. With the advent of non-biased high-throughput functional genomics such as genome-wide deletion mutant screening, novel aspects of CK2 functionality have been revealed. Our recent discoveries using the model organism Saccharomyces cerevisiae and mammalian cells demonstrate that CK2 regulates metal toxicity. Extensive literature search reveals that there are few but elegant works on the role of CK2 in regulating the sodium and zinc channels. As both CK2 and metal ions are key players in cell biology and oncogenesis, understanding the details of CK2’s regulation of metal ion homeostasis has a direct bearing on cancer research. In this review, we aim to garner the recent data and gain insights into the role of CK2 in metal ion transport

125th anniversary review : the role of proteins in beer redox stability

Flavour stability is usually approached through inhibition of reactive oxygen species (ROS). It may be possible to suppress ROS, but never to entirely eliminate them in packaged beer. The role of proteins in ROS suppression seems to have been lost in the compliant acquiescence to supply haze-free bright beer. Proteomics allows beer polypeptides to be finely resolved, identified and correlated with beer quality and stability. This has already produced a broader view of what stabilizes beer foam. No doubt it could do the same for beer stability and the broader roles that proteins, such as LTP1, can have in redox reactions and free radical suppression. Cysteine oxidation and reversibility is central to cellular signalling in biological systems. Thiol chemistry is also integral to beer redox stability. We can, and should, extrapolate the recent biological findings to the simple pleasure of creating a high-quality beer

Proteolytic extraction enhances specific detection of the novel 'S'-type low molecular weight glutenin subunit in wheat by monoclonal antibody

Specificity of monoclonal antibodies (MAbs) is required in diagnostic immunoassays. However, structural homology between a target antigen and other cellular proteins often causes promiscuous cross-reactivity of a MAb to proteins other than its target antigen, thereby hindering specificity. It is proposed in this study that specific proteolytic enzymes could be useful in the reduction or elimination of cross-reactive epitopes while retaining target epitopes for certain MAbs. The hypothesis was tested using a novel 'S'-type low molecular weight glutenin subunit (LMW-GS-'S') and its antibodies. The results demonstrated that the protease, chymotrypsin, markedly reduced the cross-reactivity in samples that would otherwise obscure the specific detection of LMW-GS-'S' by MAb F8-14E6 in an enzyme-linked immunosorbent assay (ELISA). Further investigations revealed that the working mechanism for this proteolytic treatment was indeed due to the selective cleavage that destroyed cross-reactive epitopes whilst retaining the intact specific epitope. As more proteases are being discovered or engineered, the successful utilization of protease in immunoassays as reported here will benefit general immunodiagnostic assay development in both medicine and agriculture by targeting specific epitopes with tailored proteolytic treatment of antigens

Identification of novel serpin isoforms and serpin polymorphisms among Australian wheat cultivars

Serpins constitute a large family of related proteins, the majority of which are serine protease inhibitors (from which their name is derived). They are known to be abundantly and polymorphically expressed in individual wheat cultivars. However, to date there lacks a detailed investigation into their inter-genotypic polymorphisms in larger wheat collections. In this study, a systematic proteomic approach, combining native polyacrylamide gel electrophoresis (PAGE), SDS-PAGE, two-dimensional gel electrophoresis, tandem mass spectrometry (MS/MS) and immunoblotting, was taken to characterize serpin polymorphisms among 177 Australian and 19 foreign hexaploid as well as 6 tetraploid wheat varieties. A total of seven serpin isoforms were identified and five unique expression patterns of these serpins were clearly revealed among varieties. In particular, the novel serpin isoforms 3a and 3b were highly polymorphic in the collection of wheat cultivars and are structurally diverse from the other serpin isoforms. The findings and methodology established in this study are not only immediately applicable in wheat breeding and varietal differentiation but may also be useful for delineation of serpin function in bread or pasta making in the future

Polymorphism and pedigree analysis of β-amylase isozymes in Australian wheat

β-Amylase encoded by multiple loci on chromosomes of triticeae crop plants is a starch metabolic enzyme with a significant physiological role in flour functionality. In this study, wheat grain β-amylase isozymes were characterized using an extensive Australian wheat population and genetic breeding/mapping lines. β-amylase isozymes were fractionated by high-resolution native polyacrylamide gel electrophoresis (PAGE) and validated by zymogram and peptide sequencing using electrospray ionization tandem mass spectrometry (ESI-MS/MS). Specific antibodies were generated against the β-amylase purified through protein chromatography and used for β-amylase identification. Further characterization of wheat β-amylases was carried out with immunoblotting and 2-dimensional electrophoresis (2-DE). The enzyme was found to be polymorphic among the Australian and foreign wheat cultivars, with a total of six isozymes: two common and four varying isozymes, the latter group including one novel β-amylase isozyme of fast-mobility in native PAGE, According to pedigree analysis, the origin of this particular isozyme was from the ancestral genotype, Ciano F 67

The cytotoxicity of some phenanthroline-based antimicrobial copper(II) and ruthenium(II) complexes

The cytotoxicity of copper(II) complexes with 3,4,7,8-tetramethyl-1,10-phenanthroline (TMP) or 4,7-dipyridyl-1,10-phenanthroline (DIP) ligands and ruthenium(II) complexes coordinated with the ligand 2,9-dimethyl-1,10-phenanthroline have been investigated for their toxicity in human cells. Both copper(II) complexes were found to have similar inhibitory concentrations (IC50 ~ 2–2.5 μM). Their cytotoxicity was found to be necrotic, associated with cytoplasmic vacuolisation, rounding, detachment and lack of apoptosis-associated DNA fragmentation, in comparison to the apoptotic effects of cisplatin which demonstrate adherent cell enlargement or detachment, membrane blebbing and condensation. Antimicrobial ruthenium(II) complexes demonstrated a lower renal cytotoxicity than copper(II) complexes or cisplatin (IC50 > 60 μM). [Cu(DIP)(dach)](ClO4)2 and [Cu(TMP)(dach)](ClO4)2 (where dach = 1,2-diaminocyclohexane) induced dihydroethidium-sensitive ROS and the cytotoxicity of both TMP and DIP coordinated copper(II) complexes was mitigated by catalase, highlighting a role of H2O2 generation in their mode of action. The cytotoxicity of either copper(II) complex was not affected by coincubation with organic cation transporter (OCT) inhibitors cimetidine or disopyramide, in contrast to cisplatin, suggesting a non-OCT dependent mode of uptake for the copper(II) complexes in human cells. Coincubation with copper sulfate reduced the cytotoxicity of [Cu(TMP)(dach)](ClO4)2 (3-6X). The TMP complex induced a greater degree of G2/M accumulation and micronuclei generation than the DIP complex, possibly attributable to its greater DNA binding affinity. These results highlight the potentially low genotoxicity of copper(II) complexes coordinated with TMP or DIP and polypyridyl ruthenium(II) complexes as potential antimicrobial agents

21st century skill development in remote, on-line learning biochemistry students

BACKGROUND: 21st Century skills encompass the transdisciplinary capabilities required for future work and scholarship. These skills are articulated in graduate attributes and developed through individual subjects. With the rapid transition of on-campus classes to remote learning, questions arose as to whether the same skills translated, or whether new skills were supported. Using a biochemistry unit as a case study, gap analysis of skill development was undertaken. STUDY DESIGN: The biochemistry unit (450 students) had face-to-face components (lecture, practical, workshop, examination) which were transitioned asynchronously online. Content was delivered as recordings supported by synchronous, optional online drop-in sessions held 1-3 times a week. Through staff reflection and student feedback, supported skills were identified in the two modes of delivery using a published rubric. OUTCOME: The “drop-in sessions” had participation of approximately 50 engaged students, but with the lack of compulsory laboratory sessions staff perceived many students lost reflective capability and identity to the cohort, discipline and the institution. The lack of laboratories may have reduced practical skill development and awareness of how these skills contribute to employability. However, remote delivery reinforced key 21st Century skills promoting self-direction, time management, application of professional knowledge and resilience

Comparative analyses of cytotoxicity and molecular mechanisms between platinum metallointercalators and cisplatin

Platinum(ii) metallointercalators of the type [Pt(IL)(AL)]2+, such as [(5,6-dimethyl-1,10-phenanthroline)(1S,2S- diaminocyclohexane)platinum(ii)]2+ (56MESS), are structurally different from cisplatin. This study, using a comparative transcriptomics approach, uncovered genomic expression patterns and molecular pathways that distinctively differentiated 56MESS and cisplatin in the eukaryote model organism Saccharomyces cerevisiae (yeast). Down-regulation of sulfur assimilation, cellular respiration, and energy metabolism were characteristics of 56MESS while up-regulation of these pathways and genes in cell cycle was the action of cisplatin. Furthermore, de novo purine biosynthesis and glycine metabolism were induced by 56MESS but suppressed by cisplatin. Different effects on intracellular concentrations of iron and copper were evident, with 56MESS more profoundly inducing genes controlling uptake of these ions than cisplatin. Finally, apart from 56MESS, additional metallointercalators including 56MEEN, 5MERR and 5MESS were subsequently identified to be more active in a cisplatin-resistant mouse leukaemia L1210cisR cell line than cisplatin, which provides multiple lead compounds for future drug development