Structural and functional studies of Candida albicans ALS adhesins

A great number of microorganisms are allowed to live in our bodies, mostly in our digestive and urinary systems. The vast majority of them are bacterial species. In exchange for food and a niche to grow, they break down some of the food we can't digest and synthesise essential compounds we don't produce, such as vitamins. They also provide a barrier against pathogens, as they train our immune system to fence them off and outgrow them in the competition for resources. Some fungal species can as well colonise different parts of the body and live harmlessly as commensals. However, we know that under certain conditions (especially those that affect the immune system) the natural balance of the human flora is broken and this allows organisms like fungi to overgrow and sometimes cause disease. Certain molecules are required to establish a successful interaction between hosts and these microorganisms. Many of these molecules are proteins (incidentally, they are the major component of any organism, they control our internal metabolic processes and also serve to 'scaffold' much of the shape of our bodies). The fungus Candida albicans is very frequently found in our bodies and it uses specific proteins on its surface to interact with our different tissues. Just like the machines we see everyday, proteins have a specific shape associated to their function. There is a specific class of proteins called chaperones, and their function is to help other proteins to maintain their shape, so they can carry on doing their own work. They attach firmly to 'deformed' or floppy parts of a target protein and with the help of external energy they slowly release it, as they recover their original form. As a nifty trick, Candida has managed to put one of these chaperones (or a protein with a chaperone-like activity) on its surface and create a new way to bind to tissues in humans and several animals. Interestingly, this protein doesn't seem to have the release mechanism displayed by the standard chaperones, which ultimately favours Candida, as it provides an efficient mechanism to look for proteins (with floppy regions) attached to an animal surface and then anchor to them permanently. The aim of this project is to use techniques that piece together the amino acids that form this 'chaperone-like adhesin' and accurately describe its shape. Knowing this structure will equip us to understand its function at the atomic level. As this is one of the tools that Candida uses for a successful association with its host, we will know more about how we interact with the microorganisms normally found in our gut, skin and other body sites.

The ALS family of surface glycoproteins are fundamental to the adhesive properties and morphogenesis of Candida albicans. Genome-wide inspection has identified eight different variants with homologues in other Candida spp. Their expression is stage-specific and also regulated by growth conditions. Initial characterisation of their binding properties has shown a broad range of targets in host cell surfaces and the extracellular matrix. Additionally, ALS's are determinant in the formation of biofilms, which allows C. albicans to adhere to a variety of biological and non-biological surfaces. Recent work shows that these proteins are able to bind small peptides and unstructured regions in folded domains, a function reminiscent to that of chaperones like hsp70. These observations pose an apparent contradiction: what do these proteins recognise? a well defined three-dimensional/sequential motif or simply unstructured polypeptides, e.g. free in solution or as a flexible region in an otherwise folded protein? Our preliminary data show that NT-ALS1 binds polypeptides with high affinity and broad specificity. A number of attempts by other groups to crystallise this domain have been unsuccessful, a fact likely related to the dynamic nature of the system: our NMR experiments show that the free NT-ALS1 displays alternative forms that converge on a single conformation in the protein/ligand complex. To our knowledge, this is the first example of a chaperone-like activity linked to a mechanism of adhesion and represents a novel system of molecular recognition. Importantly, the proposed activity would confer C. albicans a unique mode of binding, as it relieves the need of specific surface complementarity as observed in most ligand/receptor interactions. The structural determination of the conformations of the free protein and protein-ligand complexes is necessary to understand the mechanism that underlies the function of these adhesins.