Novel Engineered SARS-CoV-2 HR1 Trimer Exhibits Improved Potency and Broad-Spectrum Activity against SARS-CoV-2 and Its Variants.

The ongoing pandemic of COVID-19, caused by SARS-CoV-2, has substantially increased the risk to global public health. Multiple vaccines and neutralizing antibodies (nAbs) have been authorized for preventing and treating SARS-CoV-2 infection. However, the emergence and spread of the viral variants may limit the effectiveness of these vaccines and antibodies. Fusion inhibitors targeting the HR1 domain of the viral S protein have been shown to broadly inhibit SARS-CoV-2 and its variants. In theory, peptide inhibitors targeting the HR2 domain of the S protein should also be able to inhibit viral infection. However, previously reported HR1-derived peptide inhibitors targeting the HR2 domain exhibit poor inhibitory activities. Here, we engineered a novel HR1 trimer (HR1MFd) by conjugating the trimerization motif foldon to the C terminus of the HR1-derived peptide. HR1MFd showed significantly improved inhibitory activity against SARS-CoV-2, SARS-CoV-2 variants of concern (VOCs), SARS-CoV, and MERS-CoV. Mechanistically, HR1MFd possesses markedly increased α-helicity, thermostability, higher HR2 domain binding affinity, and better inhibition of S protein-mediated cell-cell fusion compared to the HR1 peptide. Therefore, HR1MFd lays the foundation to develop HR1-based fusion inhibitors against SARS-CoV-2. IMPORTANCE Peptides derived from the SARS-CoV-2 HR1 region are generally poor inhibitors. Here, we constructed a trimeric peptide HR1MFd by fusing the trimerization motif foldon to the C terminus of the HR1 peptide. HR1MFd was highly effective in blocking transductions by SARS-CoV-2, SARS-CoV-2 variants, SARS-CoV, and MERS-CoV pseudoviruses. In comparison with HR1M, HR1MFd adopted a much higher helical conformation, better thermostability, increased affinity to the viral HR2 domain, and better inhibition of S protein-mediated cell-cell fusion. Overall, HR1MFd provides the information to develop effective HR1-derived peptides as fusion inhibitors against SARS-CoV-2 and its variants.

Efficient bio-production of glutaric acid by a metabolically engineered Escherichia coli LQ-1 based on a novel nitrogen source feeding strategy.

As an important five-carbon platform chemical to synthesize polyesters and polyamides, glutaric acid is widely used in numerous biochemical fields such as consumer goods, textile, and footwear industries. However, the application of glutaric acid is limited by the low yield of its bio-production. In this study, a metabolically engineered Escherichia coli LQ-1 based on 5-aminovalerate (AMV) pathway was used for glutaric acid fed-batch fermentation. Given the significance of nitrogen source in the bio-production of glutaric acid by AMV pathway, a novel nitrogen source feeding strategy feedbacked by real-time physiological parameters was proposed after evaluating the effects of nitrogen source feeding (such as ammonia and ammonium sulfate) on glutaric acid bio-production. Under the proposed nitrogen source feeding strategy, a significantly improved glutaric acid production of 53.7 g L-1 was achieved in a 30 L fed-batch fermentation by the metabolically engineered E. coli LQ-1, which was an improvement of 52.1% over pre-optimization. Additionally, a higher conversion rate of 0.64 mol mol-1 (glutaric acid/glucose) was obtained compared with the previously reported bio-production of glutaric acid with E. coli. These results indicated that the nitrogen source feeding strategy proposed in this study will be useful for achieving the efficient and sustainable bio-based production of glutaric acid.

Rapid Optical Biosensing of SARS-CoV-2 Spike Proteins in Artificial Samples.

Tests for SARS-CoV-2 are crucial for the mass surveillance of the incidence of infection. The long waiting time for classic nucleic acid test results highlights the importance of developing alternative rapid biosensing methods. Herein, we propose a fiber-optic biolayer interferometry-based biosensor (FO-BLI) to detect SARS-CoV-2 spike proteins, extracellular domain (ECD), and receptor-binding domain (RBD) in artificial samples in 13 min. The FO-BLI biosensor utilized an antibody pair to capture and detect the spike proteins. The secondary antibody conjugated with horseradish peroxidase (HRP) reacted with the enzyme substrate for signal amplification. Two types of substrates, 3,3'-diaminobenzidine (DAB) and an advanced 3-Amino-9-ethylcarbazole (i.e., AMEC), were applied to evaluate their capabilities in enhancing signals and reaching high sensitivity. After careful comparison, the AMEC-based FO-BLI biosensor showed better assay performance, which detected ECD at a concentration of 32-720 pM and RBD of 12.5-400 pM in artificial saliva and serum, respectively. The limit of detection (LoD) for SARS-CoV-2 ECD and RBD was defined to be 36 pM and 12.5 pM, respectively. Morphology of the metal precipitates generated by the AMEC-HRP reaction in the fiber tips was observed using field emission scanning electron microscopy (SEM). Collectively, the developed FO-BLI biosensor has the potential to rapidly detect SARS-CoV-2 antigens and provide guidance for "sample-collect and result-out on-site" mode.

IgG Fc-binding motif-conjugated HIV-1 fusion inhibitor exhibits improved potency and in vivo half-life: Potential application in combination with broad neutralizing antibodies.

The clinical application of conventional peptide drugs, such as the HIV-1 fusion inhibitor enfuvirtide, is limited by their short half-life in vivo. To overcome this limitation, we developed a new strategy to extend the in vivo half-life of a short HIV-1 fusion inhibitory peptide, CP24, by fusing it with the human IgG Fc-binding peptide (IBP). The newly engineered peptide IBP-CP24 exhibited potent and broad anti-HIV-1 activity with IC50 values ranging from 0.2 to 173.7 nM for inhibiting a broad spectrum of HIV-1 strains with different subtypes and tropisms, including those resistant to enfuvirtide. Most importantly, its half-life in the plasma of rhesus monkeys was 46.1 h, about 26- and 14-fold longer than that of CP24 (t1/2 = 1.7 h) and enfuvirtide (t1/2 = 3 h), respectively. IBP-CP24 intravenously administered in rhesus monkeys could not induce significant IBP-CP24-specific antibody response and it showed no obvious in vitro or in vivo toxicity. In the prophylactic study, humanized mice pretreated with IBP-CP24 were protected from HIV-1 infection. As a therapeutic treatment, coadministration of IBP-CP24 and normal human IgG to humanized mice with chronic HIV-1 infection resulted in a significant decrease of plasma viremia. Combining IBP-CP24 with a broad neutralizing antibody (bNAb) targeting CD4-binding site (CD4bs) in gp120 or a membrane proximal external region (MPER) in gp41 exhibited synergistic effect, resulting in significant dose-reduction of the bNAb and IBP-CP24. These results suggest that IBP-CP24 has the potential to be further developed as a new HIV-1 fusion inhibitor-based, long-acting anti-HIV drug that can be used alone or in combination with a bNAb for treatment and prevention of HIV-1 infection.

Wavelength-switchable ultra-narrow linewidth fiber laser enabled by a figure-8 compound-ring-cavity filter and a polarization-managed four-channel filter.

We propose and demonstrate a high-performance wavelength-switchable erbium-doped fiber laser (EDFL), enabled by a figure-8 compound-ring-cavity (F8-CRC) filter for single-longitudinal-mode (SLM) selection and a polarization-managed four-channel filter (PM-FCF) for defining four lasing wavelengths. We introduce a novel methodology utilizing signal-flow graph combined with Mason's rule to analyze a CRC filter in general and apply it to obtain the important design parameters for the F8-CRC used in this paper. By combining the functions of the F8-CRC filter and the PM-FCF assisted by the enhanced polarization hole-burning and polarization dependent loss, we achieve the EDFL with fifteen lasing states, including four single-, six dual-, four tri- and one quad-wavelength lasing operations. In particular, all the four single-wavelength operations are in stable SLM oscillation, typically with a linewidth of <600 Hz, a RIN of ≤-154.58 dB/Hz@≥3 MHz and an output power fluctuation of ≤±3.45%. In addition, all the six dual-wavelength operations have very similar performances, with the performance parameters close to those of the four single-wavelength operations, superior to our previous work and others' similar work significantly. Finally, we achieve the wavelength-spacing tuning of dual-wavelength operations for photonic generation of tunable microwave signals, and successfully obtain a signal at 23.10 GHz as a demonstration.

Nonneutralizing Antibodies Induced by the HIV-1 gp41 NHR Domain Gain Neutralizing Activity in the Presence of the HIV Fusion Inhibitor Enfuvirtide: a Potential Therapeutic Vaccine Strategy.

A key barrier against developing preventive and therapeutic human immunodeficiency virus (HIV) vaccines is the inability of viral envelope glycoproteins to elicit broad and potent neutralizing antibodies. However, in the presence of fusion inhibitor enfuvirtide, we show that the nonneutralizing antibodies induced by the HIV type 1 (HIV-1) gp41 N-terminal heptad repeat (NHR) domain (N63) exhibit potent and broad neutralizing activity against laboratory-adapted HIV-1 strains, including the drug-resistant variants, and primary HIV-1 isolates with different subtypes, suggesting the potential of developing gp41-targeted HIV therapeutic vaccines.

An enhanced broad-spectrum peptide inhibits Omicron variants in vivo.

The continual emergence of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) variants of concern (VOCs) poses a major challenge to vaccines and antiviral therapeutics due to their extensive evasion of immunity. Aiming to develop potent and broad-spectrum anticoronavirus inhibitors, we generated A1-(GGGGS)7-HR2m (A1L35HR2m) by introducing an angiotensin-converting enzyme 2 (ACE2)-derived peptide A1 to the N terminus of the viral HR2-derived peptide HR2m through a long flexible linker, which showed significantly improved antiviral activity. Further cholesterol (Chol) modification at the C terminus of A1L35HR2m greatly enhanced the inhibitory activities against SARS-CoV-2, SARS-CoV-2 VOCs, SARS-CoV, and Middle East respiratory syndrome coronavirus (MERS-CoV) pseudoviruses, with IC50 values ranging from 0.16 to 5.53 nM. A1L35HR2m-Chol also potently inhibits spike-protein-mediated cell-cell fusion and the replication of authentic Omicron BA.2.12.1, BA.5, and EG.5.1. Importantly, A1L35HR2m-Chol distributed widely in respiratory tract tissue and had a long half-life (>10 h) in vivo. Intranasal administration of A1L35HR2m-Chol to K18-hACE2 transgenic mice potently inhibited Omicron BA.5 and EG.5.1 infection both prophylactically and therapeutically.

An amphipathic peptide targeting the gp41 cytoplasmic tail kills HIV-1 virions and infected cells.

HIV-associated morbidity and mortality have markedly declined because of combinational antiretroviral therapy, but HIV readily mutates to develop drug resistance. Developing antivirals against previously undefined targets is essential to treat existing drug-resistant HIV strains. Some peptides derived from HIV-1 envelope glycoprotein (Env, gp120-gp41) have been shown to be effective in inhibiting HIV-1 infection. Therefore, we screened a peptide library from HIV-1 Env and identified a peptide from the cytoplasmic region, designated F9170, able to effectively inactivate HIV-1 virions and induce necrosis of HIV-1-infected cells, and reactivated latently infected cells. F9170 specifically targeted the conserved cytoplasmic tail of HIV-1 Env and effectively disrupted the integrity of the viral membrane. Short-term monoadministration of F9170 controlled viral loads to below the limit of detection in chronically SHIV-infected macaques. F9170 can enter the brain and lymph nodes, anatomic reservoirs for HIV latency. Therefore, F9170 shows promise as a drug candidate for HIV treatment.

Receptor binding domain based HIV vaccines.

This paper analyzes the main trend of the development of acquired immunodeficiency syndrome (AIDS) vaccines in recent years. Designing an HIV-1 vaccine that provides robust protection from HIV-1 infection remains a challenge despite many years of effort. Therefore, we describe the receptor binding domain of gp120 as a target for developing AIDS vaccines. And we recommend some measures that could induce efficiently and produce cross-reactive neutralizing antibodies with high binding affinity. Those measures may offer a new way of the research and development of the potent and broad AIDS vaccines.