Browsing by Author "Vartema, Linda"

Bacterial surface layers (S-layers) usually consist of a single protein or glycoprotein species and they form the outermost layer of many bacteria. Three S-layer protein genes have been found in Lactobacillus brevis ATCC 14869, but the expression of only two of them, slpB and slpD, has been described in planktonic conditions. Preliminary results have suggested that the third gene, slpC, may be expressed in biofilm cells. The aim of this thesis was to study the expression of S-layer protein genes slpB, slpC and slpD in the different growth modes of L. brevis ATCC 14869. Surface proteins were detached both chemically for SDS-PAGE analysis and enzymatically for surfaceome analysis by LC-MS/MS and label-free quantification (LFQ). Messenger RNA transcript levels of the S-layer protein genes were analyzed by reverse transcriptase droplet digital PCR (RT-ddPCR). The expression of all three S-layer protein genes slpB, slpC and slpD was confirmed in the study. In the surfaceome analysis, SlpD was the most abundant protein detected on the cell surface, representing more than a third of all identified proteins, whereas SlpB and SlpC were less abundant. The S-layer proteins were detected by gel electrophoresis. Transcripts of all three genes were observed; slpD transcript amounts were the highest, whereas slpB and slpC transcript amounts were lower. The S-layer proteins and their transcripts were present in equal amounts in biofilm and planktonic cells. In this study, SlpC expression is demonstrated for the first time in L. brevis ATCC 14869. The expression of slpB, slpC and slpD was at the same level in biofilm and planktonic cells.