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.2022 Sep 26:13:1017978.
doi: 10.3389/fmicb.2022.1017978. eCollection 2022.

Phylogenetic survey of the subtilase family and a data-mining-based search for new subtilisins fromBacillaceae

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Phylogenetic survey of the subtilase family and a data-mining-based search for new subtilisins fromBacillaceae

Fabian Falkenberg et al. Front Microbiol..

Abstract

The subtilase family (S8), a member of the clan SB of serine proteases are ubiquitous in all kingdoms of life and fulfil different physiological functions. Subtilases are divided in several groups and especially subtilisins are of interest as they are used in various industrial sectors. Therefore, we searched for new subtilisin sequences of the familyBacillaceae using a data mining approach. The obtained 1,400 sequences were phylogenetically classified in the context of the subtilase family. This required an updated comprehensive overview of the different groups within this family. To fill this gap, we conducted a phylogenetic survey of the S8 family with characterised holotypes derived from the MEROPS database. The analysis revealed the presence of eight previously uncharacterised groups and 13 subgroups within the S8 family. The sequences that emerged from the data mining with the set filter parameters were mainly assigned to the subtilisin subgroups of true subtilisins, high-alkaline subtilisins, and phylogenetically intermediate subtilisins and represent an excellent source for new subtilisin candidates.

Keywords: Bacillaceae; S8 protease family; data mining; phylogenetic analysis; subtilase; subtilisin.

Copyright © 2022 Falkenberg, Bott, Bongaerts and Siegert.

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Conflict of interest statement

The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest.

Figures

FIGURE 1
FIGURE 1
Workflow of used data and methods.
FIGURE 2
FIGURE 2
MEROPS S8 holotype phylogenetic tree. Phylogenetic relationship of S8 groups and subgroups using the mature protease sequences. The tree was constructed with IQ-TREE by employing the maximum likelihood method with ultrafast bootstrap support (model: LG + I + G4, predicted by Modelfinder, 1,000 replicates). The coloured range within the labels represents the subgroups as shown within the legend. The circles around the phylogenetic tree represent the class or phylum from which the holotype originates, marked by different colours. The kexin proteases S8B were selected as the outgroup. The outer ring represents the group classification after Siezen and Leunissen (1997). For each clade, the numbers above the branches indicate the bootstrap values based on 1,000 repetitions. The tree can be accessed under the following link:https://itol.embl.de/shared/2H14VxXLj30E2.
FIGURE 3
FIGURE 3
Phylogenetic tree of 120 novel subtilisins fromBacillaceae identified by a database search. The tree was constructed with IQ-TREE using the maximum likelihood method with ultrafast bootstrap support (model: LG + I + G4 predicted by Modelfinder, 1,000 replicates). The coloured area inside and outside the labels represents the subgroups, as indicated in the legend. The theoretical isoelectric point is given for each sequence in the outer circle. Aerolysin was chosen as the outgroup. Archaea subtilisins, extremophilic subtilisins, EPR – and PopC subgroup were added as additional holotypes. All holotypes are highlighted with a yellow text colour. The bar graphs represent the cluster size of the data mining search. For each group, the numbers above the branches indicate the bootstrap values based on 1000 replications. The tree can be accessed at the following link:https://itol.embl.de/shared/2H14VxXLj30E2.
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