Comparative Structure Analysis of the Multi-Domain, Cell Envelope Proteases of Lactic Acid Bacteria

Abstract

Lactic acid bacteria (LAB) have an extracellular proteolytic system that includes a multi-domain, cell envelope protease (CEP) with a subtilisin homologous protease domain. These CEPs have different proteolytic activities despite having similar protein sequences. Structural characterization has previously been limited to CEP homologs of dairy- and human-derived LAB strains, excluding CEPs of plant-derived LAB strains. CEP structures are a challenge to determine experimentally due to their large size and attachment to the cell envelope. This study aims to clarify the prevalence and structural diversity of CEPs by using the structure prediction software AlphaFold 2. Domain boundaries are clarified based on a comparative analysis of 21 three-dimensional structures, revealing novel domain architectures of CEP homologs that are not necessarily restricted to specific LAB species or ecological niches. The C-terminal flanking region of the protease domain is divided into fibronectin type-III-like domains with various structural traits. The analysis also emphasizes the existence of two distinct domains for cell envelope attachment that are preceded by an intrinsically disordered cell wall spanning domain. The domain variants and their combinations provide CEPs with different stability, proteolytic activity, and potentially adhesive properties, making CEPs targets for steering proteolytic activity with relevance for both food development and human health.

Microbial signatures and enterotype clusters in fattening pigs: implications for nitrogen utilization efficiency

“…The present study explored L. amylovorus (ASV752) and S. alactolyticus (ASV5126) as the ASVs that contributed most to the dissimilarity of clusters LACTO and CSST. The evaluation of the proteolytic ability of LAB group members was of central interest in previous studies ( Liu et al, 2010 ; Kieliszek et al, 2021 ; Christensen et al, 2023 ). However, it was revealed that the proteolytic ability can vary widely ( Liu et al, 2010 ; Christensen et al, 2023 ).…”

“…The evaluation of the proteolytic ability of LAB group members was of central interest in previous studies ( Liu et al, 2010 ; Kieliszek et al, 2021 ; Christensen et al, 2023 ). However, it was revealed that the proteolytic ability can vary widely ( Liu et al, 2010 ; Christensen et al, 2023 ). Although all members are equipped with a proteolytic system consisting of proteinases, transport proteins and peptidases, there are inter-species differences ( Kieliszek et al, 2021 ).…”

The Microbial Community of Natural Whey Starter: Why Is It a Driver for the Production of the Most Famous Italian Long-Ripened Cheeses?

“…The contribution of the cheese microbiota to flavor development characterizes the quality and recognition of the cheese and is thus of critical significance. The final cheese flavor, as with many of the final characteristics of cheese, is due to the interactions between the cheese microorganisms, the growth substrates, proteins in the milk, and the cheese environment [14,16]. During cheese manufacturing, environmental parameters such as temperature, pH, osmolarity, and lactose concentration change significantly.…”

“…It is well established that cell lysis is a key event for the release of cytoplasmic enzymes into the cheese matrix and is crucial for understanding the contribution of different microbial cells to cheese ripening [17,[20][21][22][23][24][25][26][27][28][29][30]. In this way, the proteolysis of caseins should be considered a crucial event in determining the outcome of the process [1,7,16,31].…”