The Nanomechanical Properties of Lactococcus lactis Pili Are Conditioned by the Polymerized Backbone Pilin

Abstract

Pili produced by Lactococcus lactis subsp. lactis are putative linear structures consisting of repetitive subunits of the major pilin PilB that forms the backbone, pilin PilA situated at the distal end of the pilus, and an anchoring pilin PilC that tethers the pilus to the peptidoglycan. We determined the nanomechanical properties of pili using optical-tweezers force spectroscopy. Single pili were exposed to optical forces that yielded force-versus-extension spectra fitted using the Worm-Like Chain model. Native pili subjected to a force of 0–200 pN exhibit an inextensible, but highly flexible ultrastructure, reflected by their short persistence length. We tested a panel of derived strains to understand the functional role of the different pilins. First, we found that both the major pilin PilB and sortase C organize the backbone into a full-length organelle and dictate the nanomechanical properties of the pili. Second, we found that both PilA tip pilin and PilC anchoring pilin were not essential for the nanomechanical properties of pili. However, PilC maintains the pilus on the bacterial surface and may play a crucial role in the adhesion- and biofilm-forming properties of L. lactis.

The Role of Glycans in Bacterial Adhesion to Mucosal Surfaces: How Can Single-Molecule Techniques Advance Our Understanding?

“…Atomic force microscopy (AFM) was first developed in 1986 [ 34 ] and relies on the control of the force acting between a sharp tip, usually made of silicon nitride, and the surface while scanning a sample ( Figure 1 a). Especially in microbiology, its high-resolution imaging capacities have attracted many scientists who have used this technology to image the structure and ultrastructure of various types of bacterial cells [ 33 , 35 , 36 , 37 ].…”

“…This pili structure was also proposed for pili from Streptococcus pneumoniae TIGR4 [ 101 , 102 ] and further evidenced by force-measuring optical tweezers experiments [ 103 ]. Gram-positive pili, and particularly streptococcal pili (including lactococci) seem to share a common quaternary structure with a linear assembly of subunit that are not fully optimized to dissipate energy from environmental forces [ 18 , 34 , 103 ] as it is the case for E. coli pili [ 104 , 105 ]. Interestingly, pathogenic piliated E. coli strains express helix-like pili that dampen the external forces and minimize the load at the tip in interaction with a specific host receptor [ 105 , 106 , 107 , 108 ].…”

“…When retracting the stage, the pili are extended and unfolded until one breaks, then another one until the end of the retraction where a single pilus remains, probably the longer one. Adapted with permissions from [ 34 ]. ( d ) Force diagram corresponding to the experiment represented in panel ( c ), showing the force-extension curve of F1C pili during their unfolding (force plateaus) in a step-like manner until one single pilus is reached.…”

Surface Proteins of Lactococcus lactis: Bacterial Resources for Muco-adhesion in the Gastrointestinal Tract

“…However, under shear flow, a more critical contribution of MUB protein than pili was found. The importance of pili was further dissected in relation with their nanomechanical properties as probed with optical tweezer ( Castelain et al, 2016a , b ). AFM blocking assays also revealed that mucin neutral oligosaccharides were involved in adhesion of L. lactis TIL448 to PGM ( Le et al, 2013 ).…”

Cell Surface Properties of Lactococcus lactis Reveal Milk Protein Binding Specifically Evolved in Dairy Isolates

“…The cell surface itself is characterized by properties like charge and hydrophobicity (Ly et al, 2008 ), which are determined by the molecular composition of the cell wall. The cell wall consists of peptidoglycan (Chapot-Chartier and Kulakauskas, 2014 ), polysaccharides (Ruas-Madiedo et al, 2002 ), proteins, teichoic, and lipoteichoic acids, lipids (Pelletier et al, 1997 ) and can be decorated with a sugar pellicle (Chapot-Chartier et al, 2010 ), pili (Telford et al, 2006 ; Oxaran et al, 2012 ; Meyrand et al, 2013 ; Castelain et al, 2016 ), and/or an S-layer (van der Mei et al, 2003 ). The charge of the cell surface is determined by positively and negatively charged groups on teichoic and lipoteichoic acids, polysaccharides, proteins, and pili (Delcour et al, 1999 ; Boonaert and Rouxhet, 2000 ; Chapot-Chartier et al, 2010 ), while its hydrophobicity is related to the presence of polysaccharides, LPS, and (glyco-) proteinaceous material (Pelletier et al, 1997 ; Firoozmand and Rousseau, 2016 ), as well as pili (Tarazanova et al, 2016 ).…”