Biology subjects

Gurung, I.

Publications and source records attributed to Gurung, I..

4 recordsLinked to original sources

bioRxiv2022

Full structure/function analysis of all the pilin subunits in a type 4 pilus: a complex of minor pilins in Streptococcus sanguinis mediates binding to glycans

Type 4 filaments (T4F) - of which type 4 pili (T4P) are the archetype - are a superfamily of filamentous nanomachines nearly ubiquitous in prokaryotes. T4F are polymers of one major pilin that also contain minor pilins whose roles are often poorly understood. Here, we complete the structure/function analysis of the full set of T4P pilins in the opportunistic pathogen Streptococcus sanguinis. We determined the structure of the minor pilin PilA, which is unexpectedly similar to one of the subunits of a tip-located complex of four minor pilins, widely conserved in T4F. We found that PilA interacts and dramatically stabilises the minor pilin PilC. We determined the structure of PilC, showing that it is a modular pilin with a lectin module binding a specific subset of glycans prevalent in the human glycome, the host of S. sanguinis. Altogether, our findings support a model whereby the minor pilins in S. sanguinis T4P form a tip-located complex promoting adhesion to various host receptors. Our findings have general implications for a group of minor pilins widely conserved in T4F.

microbiology↗

bioRxiv2021

PilB from Streptococcus sanguinis is a bimodular type IV pilin with a direct role in adhesion

Type IV pili (T4P) are functionally versatile filamentous nanomachines, nearly ubiquitous in prokaryotes. They are predominantly polymers of one major pilin, but also contain minor pilins whose functions are often poorly defined, and likely to be diverse. Here, we show that the minor pilin PilB from the T4P of S. sanguinis displays an unusual bimodular 3D structure, with a bulky von Willebrand factor A-like (vWA) module "grafted" onto a small pilin module via a short unstructured loop. Structural modelling suggests that PilB is only compatible with a localisation at the tip of T4P. By performing a detailed functional analysis, we found that (i) the vWA module contains a canonical metal ion-dependent adhesion site (MIDAS), preferentially binding Mg2+ and Mn2+, (ii) abolishing metal-binding has no impact on the structure of PilB or piliation, (iii) metal-binding is important for S. sanguinis T4P-mediated twitching motility and adhesion to eukaryotic cells, and (iv) the vWA module shows an intrinsic binding ability to several host proteins. These findings reveal an elegant, yet simple, evolutionary tinkering strategy to increase T4P functional versatility, by grafting an adhesive module onto a pilin for presentation by the filaments. This strategy appears to have been extensively used by bacteria, in which modular pilins are widespread and exhibit an astonishing variety of architectures.

microbiology↗

bioRxiv2020

Construction of a complete set of Neisseria meningitidis defined mutants - the NeMeSys collection - and its use for the phenotypic profiling of the genome of an important human pathogen

One of the great challenges in biology is to determine the function of millions of genes of unknown function. Even in model bacterial species, there is a sizeable proportion of such genes, which has fundamental and practical consequences. Here, we constructed a complete collection of defined mutants in protein-coding genes - named NeMeSys 2.0 - in the human pathogen Neisseria meningitidis, consisting of individual mutants in 1,584 non-essential genes. This effort identified 391 essential genes - broadly conserved in other bacteria - leading to a full overview of the essential meningococcal genome, associated with just four underlying basic biological functions: 1) expression of genome information, 2) preservation of genome information, 3) cell membrane structure/function, and 4) cytosolic metabolism. Subsequently, we illustrated the utility of the NeMeSys 2.0 collection for determining gene function by identifying 1) a novel and conserved family of histidinol-phosphatase, 2) 20 genes, including three new ones, involved in the biology of type IV pili, a widespread virulence factor, and 3) several conditionally essential genes found in regions of genome plasticity encoding antitoxins and/or immunity proteins, which become dispensable when the gene encoding the cognate toxin is deleted. These findings have implications beyond the meningococcus. The NeMeSys 2.0 collection is an invaluable resource paving the way for a global phenotypic landscape in a major human bacterial pathogen.

microbiology↗

bioRxiv2018

In depth biochemical and structural analysis of a Gram-positive type IV pilus

Type IV pili (Tfp) are functionally versatile filaments, widespread in prokaryotes, that belong to a large class of filamentous nanomachines known as type IV filaments (Tff). Although Tfp have been extensively studied in several Gram-negative pathogens where they function as key virulence factors, many aspects of their biology remain poorly understood. Here, we performed a global biochemical and structural analysis of Tfp in a recently emerged Gram-positive model, Streptococcus sanguinis. In particular, we focused on the five pilins and pilin-like proteins involved in Tfp biology in S. sanguinis. We found that the two major pilins, PilE1 and PilE2, (i) follow widely conserved principles for processing by the prepilin peptidase PilD and for assembly into filaments; (ii) display only one of the post-translational modifications frequently found in pilins, i.e. a methylated N-terminus; (iii) are found in the same hetero-polymeric filaments; and (iv) are not functionally equivalent. The 3D structure of PilE1, solved by NMR, revealed a classical pilin fold with a highly unusual flexible C-terminus. Intriguingly, PilE1 more closely resembles pseudopilins forming shorter Tff than bona fide Tfp-forming major pilins, underlining the evolutionary relatedness among different Tff. Finally, we show that S. sanguinis Tfp contain a low abundance of three additional proteins processed by PilD, the minor pilins PilA, PilB, and PilC. These findings provide the first global biochemical and structural picture of a Gram-positive Tfp and have fundamental implications for our understanding of a widespread class of filamentous nanomachines.

microbiology↗