The mannose-specific lectin domains of Flo1p fromSaccharomyces cerevisiaeand Lg-Flo1p fromS. pastorianus: crystallization and preliminary X-ray diffraction analysis of the adhesin–carbohydrate complexes

Abstract

Flo1p and Lg-Flo1p are two cell-wall adhesins belonging to the Flo (flocculation) protein family from the yeasts Saccharomyces cerevisiae and S. pastorianus. The main function of these modular proteins endowed with calcium-dependent lectin activity is to mediate cell-cell adhesion events during yeast flocculation, a process which is well known at the cellular level but still not fully characterized from a molecular perspective. Recently, structural features of the N-terminal Flo lectin domains, including the N-terminal domain of Lg-Flo1p (N-Lg-Flo1p), and their interactions with carbohydrate molecules have been investigated. However, structural data concerning the N-terminal domain of Flo1p (N-Flo1p), which is the most specific among the Flo proteins, are missing and information about the N-Lg-Flo1p-carbohydrate interaction still lacks detailed structural insight. Here, the crystallization and preliminary X-ray characterization of the apo form and the mannose complex of N-Flo1p and X-ray analysis of N-Lg-Flo1p crystals soaked in -1,2-mannobiose are reported. The N-Flo1p crystals diffracted to a resolution of 1.43 Å in the case of the apo form and to 2.12 Å resolution for the mannose complex. Both crystals were orthorhombic and belonged to space group P2 1 2 1 2 1 , with one molecule in the asymmetric unit. The N-Lg-Flo1p--1,2-mannobiose complex crystal diffracted to 1.73 Å resolution and belonged to the monoclinic space group P12 1 1 with two molecules in the asymmetric unit.

Molecular Mechanism of Flocculation Self-Recognition in Yeast and Its Role in Mating and Survival

“…The glycosylated N-terminal domain of Flo1p was purified by a combination of affinity chromatography and gel filtration after expression in S. cerevisiae ( 22 ). Expression and purification of the N-terminal domain of Flo1p and Lg-Flo1p from Escherichia coli have been described elsewhere ( 126 ) (see Text S1 in the supplemental material).…”

“…Crystallization and X-ray data collection for N-Flo1p (in apo form and in complex with mannose) and N-Lg-Flo1p (in complex with α-1,2-mannobiose) have been described elsewhere ( 126 ). The structure of the flocculin N-terminal domains (PDB codes 4LHK [N-Lg-Flo1p–mannobiose], 4LHL [apo–N-Flo1p], and 4LHN [N-Flo1p–mannose]) were solved by molecular replacement (see Text S1 and Table S1 in the supplemental material).…”

Distinctive ligand‐binding specificities of tandem PA14 biomass‐sensory elements from Clostridium thermocellum and Clostridium clariflavum

“…The Protein Data Bank was therefore searched with C. thermocellum and C. clariflavum PA14 A / PA14 B (monomers) as queries, using the DALI server, and several similar structures were found. High structural similarity of 2.2 and 2.3 Å was found with Saccharomyces cerevisiae flocculin (Flo) proteins (2XJS, 4AIL, and 4LHL‐A) and Candida glabrata epithelial adhesins (Epa) (4ASL), respectively . The RsgI_PA14 modules and the PA14 modular structures of the flocculin and adhesin proteins superimpose well at the core part of the β‐sandwich region, although the adhesin (Figure A, gray) and flocculin (Figure A, purple) proteins are decorated by additional structural regions, which are absent from the RsgI PA14 modules.…”

FLO5 gene controls flocculation phenotype and adhesive properties in a Saccharomyces cerevisiae sparkling wine strain

“…The ability to adhere to other cells and substrates is a crucial feature of microorganisms that can be used both for ‘defensive’ or ‘offensive’ purposes. In Saccharomyces cerevisiae , it has the univocal purpose of allowing yeast survival in different biological contexts 1 . Yeast strains display different adhesion phenotypes such as biofilm, invasive growth, agglutination, chain formation co-flocculation and flocculation.…”