Proteomic analysis of the periodontal pathogen Prevotella intermedia secretomes in biofilm and planktonic lifestyles

Maribasappa Karched 1, Radhika G Bhardwaj 2, Muawia Qudeimat 3, Areej Al-Khabbaz 4, Arjuna Ellepola 2

Affiliations

04 April 2022

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doi: 10.1038/s41598-022-09085-0

Abstract

Prevotella intermedia is an important species associated with periodontitis. Despite the remarkable clinical significance, little is known about the molecular basis for its virulence. The aim of this study was to characterize the secretome of P. intermedia in biofilm and planktonic life mode. The biofilm secretome showed 109 proteins while the planktonic secretome showed 136 proteins. The biofilm and the planktonic secretomes contained 17 and 33 signal-peptide bearing proteins, 13 and 18 lipoproteins, respectively. Superoxide reductase, sensor histidine kinase, C40 family peptidase, elongation factor Tu, threonine synthase etc. were unique to biofilm. Of the ~ 30 proteins with predicted virulence potential from biofilm and planktonic secretomes, only 6 were common between the two groups, implying large differences between biofilm and planktonic modes of P. intermedia. From Gene Ontology biofilm secretome displayed a markedly higher percent proteins compared to planktonic secretome in terms of cellular amino acid metabolic process, nitrogen compound metabolic process etc. Inflammatory cytokine profile analysis revealed that only the biofilm secretome, not the planktonic one, induced important cytokines such as MIP-1α/MIP-1β, IL-1β, and IL-8. In conclusion, the revealed differences in the protein profiles of P. intermedia biofilm and planktonic secretomes may trigger further questions about molecular mechanisms how this species exerts its virulence potential in the oral cavity.

Conflict of interest statement

The authors declare no competing interests.

Figures

Figure 1 Analysis of the proteome of P. intermedia biofilm and planktonic cells. (A) SDS–PAGE gel showing protein bands from protein preparations: biofilm (lane 1) and planktonic cells (lane 2). (B) Western blot analysis of the secretome preparations (lane 1 = biofilm, lane 2 = planktonic) and the WCP (lane 3) using an antibody for the cytoplasmic marker protein FtsZ. (C) Protein sequences from LC–MS analysis of the secretome were analyzed by an in silico 2DE tool.

Figure 1 Analysis of the proteome of P. intermedia biofilm and planktonic cells. (A) SDS–PAGE gel showing protein bands from protein preparations: biofilm (lane 1) and planktonic cells (lane 2). (B) Western blot analysis of the secretome preparations (lane 1 = biofilm, lane 2 = planktonic) and the WCP (lane 3) using an antibody for the cytoplasmic marker protein FtsZ. (C) Protein sequences from LC–MS analysis of the secretome were analyzed by an in silico 2DE tool.

Figure 2 Subcellular localization of proteins. Protein FASTA sequences were analyzed for their subcellular localization using the bioinformatics tool CELLO2GO. The results obtained were compared with other predictions tools such as PSORTb and SignalP.

Figure 2 Subcellular localization of proteins. Protein FASTA sequences were analyzed for their subcellular localization using the bioinformatics tool CELLO2GO. The results obtained were compared with other predictions tools such as PSORTb and SignalP.

Figure 3 Gene Ontology analysis of P. intermedia proteomes from biofilm and planktonic cells. Gene Ontology annotation was achieved using Blast2GO and an online software “CELLO2GO”. Protein sequences were grouped into 3 categories based on their properties and functions.

Figure 3 Gene Ontology analysis of P. intermedia proteomes from biofilm and planktonic cells. Gene Ontology annotation was achieved using Blast2GO and an online software “CELLO2GO”. Protein sequences were grouped into 3 categories based on their properties and functions.

Figure 4 Functional protein association networks of P. intermedia proteome from biofilm and planktonic cells. The online tool STRING was used for grouping the secreted proteins based on functional networks. Minimum interaction scores were set at a strong confidence level of 0.7. The three major network groups formed are shown in dotted circles. The thickness of the lines in the network indicates the strength of data support.

Figure 4 Functional protein association networks of P. intermedia proteome from biofilm and planktonic cells. The online tool STRING was used for grouping the secreted proteins based on functional networks. Minimum interaction scores were set at a strong confidence level of 0.7. The three major network groups formed are shown in dotted circles. The thickness of the lines in the network indicates the strength of data support.

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