Candida albicans Agglutinin-Like Sequence (Als) Family Vignettes: A Review of Als Protein Structure and Function

Abstract

Approximately two decades have passed since the description of the first gene in the Candida albicans ALS (agglutinin-like sequence) family. Since that time, much has been learned about the composition of the family and the function of its encoded cell-surface glycoproteins. Solution of the structure of the Als adhesive domain provides the opportunity to evaluate the molecular basis for protein function. This review article is formatted as a series of fundamental questions and explores the diversity of the Als proteins, as well as their role in ligand binding, aggregative effects, and attachment to abiotic surfaces. Interaction of Als proteins with each other, their functional equivalence, and the effects of protein abundance on phenotypic conclusions are also examined. Structural features of Als proteins that may facilitate invasive function are considered. Conclusions that are firmly supported by the literature are presented while highlighting areas that require additional investigation to reveal basic features of the Als proteins, their relatedness to each other, and their roles in C. albicans biology.

Keywords: Als proteins; Candida albicans; adhesion; aggregation; attachment; fungus; gene family; invasion.

FIGURE 1

Als protein structure. (A) Line drawing of a representative Als protein, using C. albicans Als3 as the example. A detailed schematic comparing the basic features of all C. albicans Als proteins was published previously (Hoyer et al., 2008). The various domains are labeled as they are discussed in this review: NT (also called NT-Als), T, TR, and CT. Early literature referred to the sequences N-terminal of the TRs as the ‘NT’; this region is indicated by the solid line below the main drawing. Als proteins include a secretory signal sequence which is processed, so absent from the mature protein. Als proteins also encode a consensus sequence for GPI (glycosyl-phosphatidylinositol) anchor addition. The GPI anchor subsequently is processed and the mature protein linked to beta-1,6-glucan in the C. albicans cell wall (Kapteyn et al., 2000). Numbering schemes found in the literature may be confusing because some start at the initial Met (shown above the line drawing) while others start at the N-terminal amino acid of the mature protein, following cleavage of the secretory signal peptide (e.g., amino acid 18 of the unprocessed sequence in many of the Als proteins; shown below the line drawing). Clarifications are provided throughout the review to indicate whether the numbering scheme arises from the unprocessed (signal sequence present) or processed (cleaved signal sequence) protein. (B) X-ray crystallographic structure of the NT domain from Als9-2 in complex with the C-terminal peptide from fibrinogen-γ (red; Salgado et al., 2011) that fits into the protein’s PBC. An invariant Lys residue (K59, using a numbering scheme for the processed protein; blue) at the end of the PBC recognizes the C-terminal carboxyl group of the peptide ligand. The overall fold of the protein involves eight conserved Cys residues that form four disulfide bonds. In the ligand-bound form of the protein, the AFR (gray) attaches to the NT-Als surface. The AFR is unattached to the NT-Als surface in protein molecules that do not have a ligand in the PBC.

FIGURE 2

Schematics of NT-Als protein structure to illustrate the location of mutations used to deduce the ligand-binding mechanism. (A) Cross-section of overall NT-Als3 structure highlighting the location of the PBC and key residues used in loss-of-function mutants. Note that the indicated mutations were introduced without altering NT-Als surface properties. Amino acid numbering reflects the processed (signal peptide removed) form of the protein (Lin et al., 2014). (B) Expanded PBC detail showing entry of a model peptide and location of amino acids included in the functional analysis using the structure of NT-Als9-2 (Salgado et al., 2011).

FIGURE 3

Models proposed to explain function of the AFR in Als protein interactions. (A) Force-induced aggregation of Als proteins on the surface of the same cell from Lipke et al. (2012). Homotypic binding between NT domains of Als proteins is proposed to trigger force required to pull apart an Als protein, exposing the AFR for interaction with AFR sequences on other Als proteins. (B) Variable conformation of the AFR in relation to the NT domain of Als3 on the C. albicans cell surface based on Lin et al. (2014). Newly synthesized Als protein can either bind ligand via the PBC, which results in the AFR attaching to the NT domain surface (left) or use its free AFR to interact with others, forming protein and cellular aggregates (right). Note that the model in (A) and the model in (B) show different artistic interpretations of AFR placement, with (B) showing an exaggerated scale of the NT portion of the molecule (especially the AFR) to emphasize those interactions. (A) Reprinted from Lipke et al. (2012), with permission from Elsevier. (B) This research was originally published in Lin et al. (2014). Reprinted with permission from The American Society for Biochemistry and Molecular Biology.

FIGURE 4

Mechanisms of NT domain interactions between purified proteins (A) and between mature, full-length Als proteins on the C. albicans cell surface (B,C). (A) Purified NT-Als proteins may interact by two mechanisms. The first involves PBC-mediated recognition of the free C-terminal peptide, leading to oligomerization of the NT-Als molecules (left). The second mechanism involves aggregation mediated by the AFR (right). Because the NT domain is a small portion of the full-length, mature Als protein, PBC-mediated oligomerization of the proteins cannot explain aggregation between Als molecules on the C. albicans cell surface. These interactions are more likely attributable to the AFR (B). The AFR of mature, full-length Als proteins can also promote Als–Als-mediated aggregation between different C. albicans cells (C).

FIGURE 5

Potential mechanisms to explain the PBC-mediated adhesive/invasive interactions of Als3 with host cells. (A) The Als3 PBC may interact with extracellular features of intact cadherins or other mammalian cell-surface proteins. (B) C. albicans may release proteases to facilitate partial digestion of cell-surface proteins, producing free C termini that are anchored to the host-cell membrane and competent for interaction with the Als3 PBC. (C) C. albicans may damage the host-cell membrane and promote translocation of Als3 into the host-cell cytoplasm where it may contact the C termini of membrane-anchored proteins.