Bacterial lipid modification of proteins for novel protein engineering applications

Functioning of proteins efficiently at the solid-liquid interface is critical to not only biological but also modern man-made systems such as ELISA, liposomes and biosensors. Anchoring hydrophilic proteins poses a major challenge in this regard. Lipid modification, N-acyl-S-diacylglyceryl-Cys, providing an N-terminal hydrophobic membrane anchor is a viable solution that bacteria have successfully evolved but remains unexploited. Based on the current understanding of this ubiquitous and unique bacterial lipid modification it is possible to use Escherichia coli, the popular recombinant protein expression host, for converting a non-lipoprotein to a lipoprotein with a hydrophobic anchor at the N-terminal end. We report two strategies applicable to non-lipoproteins (with or without signal sequences) employing minimal sequence change. Taking periplasmic Shigella apyrase as an example, its signal sequence was engineered to include a lipobox, an essential determinant for lipid modification, or its mature sequence was fused to the signal sequence of abundant outer membrane lipoprotein, Lpp. Lipid modification was proved by membrane localization, electrophoretic mobility shift and mass spectrometric analysis. Substrate specificity and specific activity measurements indicated functional integrity after modification. In conclusion, a convenient protein engineering strategy for converting non-lipoprotein to lipoprotein for commercial application has been devised and tested successfully

Mode shift behaviour and user willingness to adopt the electric two-wheeler: A study based on Indian road user preferences

As per statistics, two-wheeler (TW) alone shares the highest number of vehicle registrations in India, which develops the various transportation-related issues such as traffic conflicts, congestions, and pollutions. Electric two-wheelers (E-TW) are a better alternative to conventional two-wheelers because of their significant advantage in mitigating environmental impacts. But E-TWs are less attractive among road users due to unawareness of the benefits of E-TW. In addition, the traditional methods are less accurate in predicting users' mode choice behavior because of their limitations. Therefore, there is a need to conduct a study to understand the road user's willingness to adopt E-TW and find a suitable method for predicting mode choice behavior accurately. This study analyzes the Indian road users encouraging and discouraging factors to adopt E-TW and investigates the application of non-traditional models for estimating mode shift behaviour towards E-TW. Based on the literature review and expert opinion, a detailed questionnaire form was framed, and a total of 522 samples were collected from four states of India. The data findings show that Indian road users prefer TW compared to public transport, private four-wheeler, paratransit, and non-motorized transport because of its easy to ride, low maintenance, fast and convenient travel nature. The environmental concern of reducing air pollution and lower vehicle operating costs are significant factors that encourage E-TW adoption. However, the non-availability of charging infrastructure, lower speed, higher initial purchase cost, and lack of awareness about EVs are the significant discouraging factors in adopting E-TW in India. Further, Machine Learning (ML) methods were adopted to predict the mode shift behaviour from the fuel based TW to E-TW, and the results were compared with the Binary Logit (BL) method. The model results indicated that Support vector machine predicted the mode shift behavior with the highest accuracy rate compared with other methods such as Artificial Neural Network, K-Nearest Neighbor, Random Forest, and BL. The outcome of this study would help the transportation planner, EV manufacturers, researchers, and policymakers to understand the Indian user's preference to adopt E-TW

What influences the implementation of the New Zealand Stroke Guidelines for Physiotherapists and Occupational Therapists?

Purpose To explore perceived barriers and facilitators to the use of the New Zealand (NZ) stroke guidelines by occupational therapists and physiotherapists. Methods A qualitative descriptive methodology was used. Eligible physiotherapists and occupational therapists (NZ registered, working in one of two hospitals, treating at least 10 patients with stroke in the previous year) were invited to participate in semi-structured interviews to elicit their perceptions of the utility and feasibility of the NZ stroke guidelines and identify barriers and facilitators to their implementation. All interviews were audio-recorded and transcribed. Conventional content analysis with constant comparative methods was used for coding and analysis. Results The main themes influencing guideline implementation were resources and characteristics of the guidelines, the organization, the patient and family and the therapist. Insufficient resources were a major barrier that crossed many of the themes. Participants suggested a range of strategies relating to the organization to improve therapists’ alignment to the guidelines. Conclusion Alignment to the guidelines in NZ is influenced both positively and negatively by a range of interacting factors, consistent with other studies. Alignment might be improved by the introduction of some relatively simple strategies, such as ring-fencing time for access to resources and training in the use of the guidelines. Many of the barriers and related interventions are likely to be more complex. Implications for rehabilitation Alignment with stroke guidelines has been shown to improve patient outcomes. Therapist alignment with the implementation of the New Zealand stroke guidelines is influenced by guideline characteristics, organizational characteristics, resources, patient and family characteristics and therapist characteristics. Frequently encountered barriers related to limited resources, particularly time. Ring-fencing regular time for access to resources and training in the use of guidelines are examples of simple strategies that may reduce barriers

Biological evaluation of redox stable cisplatin/Cu(II)-DNA adducts as potential anticancer agents

<p>A series of non-enolizable β-diketonate-based copper(II) complexes with LCuCl₂ [L = Knoevenagel condensates of curcumin (Salcimine) and methylacetoacetate (SalMaA)-based Schiff bases] chromospheres as functional models of chemotherapy drug cisplatin were investigated for their covalent interaction with herring sperm DNA. The synthesis and structural characterization of <b>1a</b> and <b>1b</b> have been reported in our previous article. However, their DNA interactions and cytotoxicity properties were not studied. These analyses have been carried out mainly through electrochemical techniques supplemented with spectral, relative viscosity, gel electrophoresis techniques, and AGS cancer cells using MTT assay. The cytotoxic activities of the ligand, curcumin-based copper complex, and cisplatin were tested against the AGS cancer cell line under similar experimental conditions showing that the complex exhibited cancer cell inhibitory rate closer to cisplatin even at low concentration. This was also seen in the docking of the Cu-complex onto a rich guanine B-DNA decamer, where a Cu–N3_(guanine) interaction instead of Pt-N7 as cisplatin is detected. The obtained results in this study prove that these complexes could be a promising substitute for cisplatin as a new family of non-platinum-based anticancer metallo-drugs after <i>in vivo</i> tests on animal models.</p