The rise in antibiotic resistance has led to an increased research focus on discovery of new antibacterial candidates. While broad-spectrum antibiotics are widely pursued, there is evidence that resistance arises in part from the wide spread use of these antibiotics. Our group has developed a system to produce protein affinity agents, called synbodies, which have high affinity and specificity for their target. In this report, we describe the adaptation of this system to produce new antibacterial candidates towards a target bacterium. The system functions by screening target bacteria against an array of 10,000 random sequence peptides and, using a combination of membrane labeling and intracellular dyes, we identified peptides with target specific binding or killing functions. Binding and lytic peptides were identified in this manner and in vitro tests confirmed the activity of the lead peptides. A peptide with antibacterial activity was linked to a peptide specifically binding Staphylococcus aureus to create a synbody with increased antibacterial activity. Subsequent tests showed that this peptide could block S. aureus induced killing of HEK293 cells in a co-culture experiment. These results demonstrate the feasibility of using the synbody system to discover new antibacterial candidate agents.

Bioethics is an important aspect of the core competency of biology of understanding the relationship between science and society, but because of the controversial nature of the topics covered in bioethics courses, different groups of students may experience identity conflicts or discomfort when learning about them. However, no previous studies have investigated the impact of undergraduate bioethics students’ experiences in bioethics courses on their opinions and comfort. To fill this gap in knowledge, we investigated undergraduate bioethics students’ attitudes about and comfort when learning abortion, gene editing, and physician assisted suicide, as well as how their gender, religious, and political identity influence their attitudes and changes in their attitudes after instruction. We found that religious students were less supportive of gene editing, abortion, and physician assisted suicide than nonreligious students, non-liberal students were less supportive of abortion and physician assisted suicide than liberal students, and women were less supportive of abortion than men. Additionally, we found that religious students were less comfortable than nonreligious students when learning about gene editing, abortion, and physician assisted suicide, and non-liberal students were less comfortable than liberal students when learning about abortion. When asked how their comfort could have been improved, those who felt that their peers or instructors could have done something to increase their comfort most commonly cited that including additional unbiased materials or incorporating materials and discussions that cover both sides of every controversial issue would have helped them to feel more comfortable when learning about gene editing, abortion, and physician assisted suicide. Finally, we found that students who were less comfortable learning about abortion and physician assisted suicide were less likely to participate in discussions regarding those topics. Our findings show that students in different groups not only tend to have different support for controversial topics like gene editing, abortion, and physician assisted suicide, but they also feel differentially comfortable when learning about them, which in turn impacts their participation. We hope that this work helps instructors to recognize the importance of their students’ comfort to their learning in bioethics courses, and from this study, they can take away the knowledge that students feel their comfort could be most improved by the incorporation of additional inclusive materials and course discussions regarding the controversial topics covered in the course.

The heat-labile toxins (LT) produced by enterotoxigenic Escherichia coli display adjuvant effects to coadministered antigens, leading to enhanced production of serum antibodies. Despite extensive knowledge of the adjuvant properties of LT derivatives, including in vitro-generated non-toxic mutant forms, little is known about the capacity of these adjuvants to modulate the epitope specificity of antibodies directed against antigens. This study characterizes the role of LT and its non-toxic B subunit (LTB) in the modulation of antibody responses to a coadministered antigen, the dengue virus (DENV) envelope glycoprotein domain III (EDIII), which binds to surface receptors and mediates virus entry into host cells. In contrast to non-adjuvanted or alum-adjuvanted formulations, antibodies induced in mice immunized with LT or LTB showed enhanced virus-neutralization effects that were not ascribed to a subclass shift or antigen affinity. Nonetheless, immunosignature analyses revealed that purified LT-adjuvanted EDIII-specific antibodies display distinct epitope-binding patterns with regard to antibodies raised in mice immunized with EDIII or the alum-adjuvanted vaccine. Notably, the analyses led to the identification of a specific EDIII epitope located in the EF to FG loop, which is involved in the entry of DENV into eukaryotic cells. The present results demonstrate that LT and LTB modulate the epitope specificity of antibodies generated after immunization with coadministered antigens that, in the case of EDIII, was associated with the induction of neutralizing antibody responses. These results open perspectives for the more rational development of vaccines with enhanced protective effects against DENV infections.

Background: Chemistry and particularly enzymology at surfaces is a topic of rapidly growing interest, both in terms of its role in biological systems and its application in biocatalysis. Existing protein immobilization approaches, including noncovalent or covalent attachments to solid supports, have difficulties in controlling protein orientation, reducing nonspecific absorption and preventing protein denaturation. New strategies for enzyme immobilization are needed that allow the precise control over orientation and position and thereby provide optimized activity.

Methodology/Principal Findings: A method is presented for utilizing peptide ligands to immobilize enzymes on surfaces with improved enzyme activity and stability. The appropriate peptide ligands have been rapidly selected from high-density arrays and when desirable, the peptide sequences were further optimized by single-point variant screening to enhance both the affinity and activity of the bound enzyme. For proof of concept, the peptides that bound to β-galactosidase and optimized its activity were covalently attached to surfaces for the purpose of capturing target enzymes. Compared to conventional methods, enzymes immobilized on peptide-modified surfaces exhibited higher specific activity and stability, as well as controlled protein orientation.

Conclusions/Significance: A simple method for immobilizing enzymes through specific interactions with peptides anchored on surfaces has been developed. This approach will be applicable to the immobilization of a wide variety of enzymes on surfaces with optimized orientation, location and performance, and provides a potential mechanism for the patterned self-assembly of multiple enzymes on surfaces.