Vani Verma postdoc researcher FWO

Research Vani Verma

Research team

Expertise

Bacterial infections play a significant role in various lung diseases, including community-acquired pneumonia. To infiltrate the lungs, bacterial pathogens produce numerous proteases that are essential for functions such as cell viability, physiology, and virulence. One such example is Streptococcus pneumoniae, which relies on various virulence factors, including the serine-based high-temperature requirement A protease (HtrA). The HtrA protease activity is critical for breaking down biofilms and promoting competence, a crucial step in the development of multi-drug resistant strains. However, our understanding of the molecular mechanisms governing SP-HtrA's proteolytic activity during different infection stages remains incomplete, primarily due to the limited methods available to study HtrA protease activity. This understanding is not only essential but also timely, given the increasing multidrug resistance among pathogens. Several lines of evidence indicate that proteases, in general, play a crucial role in pathogen survival and colonization. Therefore, elucidating the previously unknown virulent roles of these proteases is vital, as it can lead to the discovery of novel therapeutic targets or approaches. Consequently, my research at the University of Antwerp is divided into two main components: The development of highly sensitive and selective activity-based probes (ABPs) through activity-based protein profiling. These ABPs will covalently modify the active site of the protease, enabling the real-time monitoring of protease cleavage in living cells and tissues. The application of these ABPs, following biochemical validation, in in vitro and in vivo infection models. This project involves various techniques, including solid-phase peptide synthesis, organic synthetic chemistry, biochemical methods, protein expression and purification, proteomics, and imaging, among others.

Development of chemical tools for deciphering the virulent roles of Streptococcus pneumoniae HtrA protease. 01/10/2024 - 30/09/2027

Abstract

Streptococcus pneumoniae (SP) is a leading cause of community-acquired pneumonia, with a concerning rise in drug-resistant strains. Central to its virulence is the high-temperature requirement A protease (HtrA), which plays a dual role as a chaperone and protease. HtrA is pivotal in SP's pathological processes, offering an innovative target for antibacterial strategies. Yet, the exact role of HtrA during infection remains elusive, posing challenges for drug development. This project aims to elucidate SP-HtrA's protease function during the infection process. We plan to develop activity-based probes (ABPs) for detecting and localizing active SP-HtrA proteases, alongside cyclic peptides as specific, irreversible inhibitors. These tools and inhibitors will contain a specific peptide scaffold, tailored to SP-HtrA's active site through advanced techniques, including positional scanning combinatorial libraries and phage display, ensuring specificity and avoiding interference with human HtrAs. We will demonstrate the power of these ABPs and inhibitors in both in vitro and in vivo infection models by offering proteomic insights into the activity status of SP-HtrA within an infection context. Overall, this project will provide a novel strategy to examine SP-HtrA protease activity during infection, which goes beyond the capabilities of conventional expression profiling technologies.

Researcher(s)

Research team(s)

Project type(s)

  • Research Project

Chemical probes for imaging bacterial proteases in lung infections. 01/11/2022 - 31/10/2024

Abstract

Bacterial infections play an important role in many lung diseases, including infective exacerbations of chronic obstructive lung disease (COPD) or community-acquired pneumonia (CAP), one of the most frequently diagnosed diseases worldwide. In order to infect the lung, bacterial pathogens produce numerous proteases with essential functions in cell viability, physiology and virulence. For example, Elastase B of Pseudomonas aeruginosa causes extensive lung damage during pneumonia. These proteases are promising candidates as both antimicrobial drug target and biomarker for lung infection. However, the precise mechanisms in which they contribute to virulence is often unclear, hampering drug and biomarker discovery. The research topic described herein uses the power of chemical probes to understand the virulent roles of bacterial proteases, for which we currently lack the tools to determine. The selected candidate will develop highly sensitive and selective chemical probes that will report on bacterial proteases activity in infection models and patient samples. This will allow to (1) uncover yet unknown virulence functions of bacterial proteases in lung diseases, such as biofilm formation and persistence and (2) evaluate bacterial proteases as potential biomarkers for bacterial lung infections. The two major groups of chemical tools to profile protease activity are activity-based probes (ABPs) and substrate probes. ABPs are small molecules that bind covalently to the active site of target enzymes. They usually contain a recognition sequence, a detection tag and an electrophilic or photoreactive group to bind into the active site. Substrate probes typically comprise of recognition sequences flanked by reagents that generate a fluorescent readout after cleavage. The candidate will synthesize such tools based on known inhibitors or substrates of bacterial proteases produced by pathogens involved in lung diseases, such as Haemophilus influenzae or Streptococcus pneumoniae. Many lung pathogens exert a particularly virulent behaviour by persisting inside epithelial cells, allowing them to evade the human immune response and antibiotic treatment. However, the role of bacterial proteases during persistence is often unknown. Thus, the candidate will apply the new probes, after biochemical validation, in in vitro and in vivo infection models to monitor the enzyme activity during persistence (collaboration with Prof. Paul Cos). Moreover, due to their high selectivity for the respective bacterial proteases, the novel tools will be perfectly suited as activity-based diagnostics. Although diagnosis is critical in acute respiratory illness, diagnostic tests that rapidly clarify the causative pathogen are often lacking and urgently needed. Therefore, the candidate will apply the probes in samples of hospitalized patients with lung infections and evaluate them as potential activity-based diagnostic tools (collaboration with Prof. Thérèse Lapperre).

Researcher(s)

Research team(s)

Project type(s)

  • Research Project