Function of your Protein
This is an , specifically Epa9, it binds to a host cell (human epithelial cell) to provide a host cell recognition to invade the host. In this case, this epithelial adhesion belongs to the fungus Candida glabrata. This fungus has more than just one Epa to create host cell recognition, it has over 20 Epa's. These proteins can adhere Candida glabrata to a human epithelial cell by specifically binding to different carbohydrates on the outside of the cell. This interaction between the Epas and the carbohydrates is what creates the adhesion for host cell recognition.
Biological relevance and broader implications
Candida glabrata is a fungus of high concern as it infects the host through the bloodstream. Unfortunately, this is a life-threatening infection for humans and upwards of 29% of all cases are life-threatening. As this does affect the human race is it of high relevance to study in health sciences. Understanding how this fungus can infect the bloodstream is needed to slow and possibly stop Candida glabrata from infecting other people. The approach in this paper is on the epithelial adhesions and altering their composition around the binding site. By altering conserved and un-conserved areas in its binding site we can better understand what hot spots are needed for good binding to the carbohydrates on the human epithelial cells. Understanding this protein's function is beneficial to understanding how other possible fungi infect hosts in a parasitic relationship. In this page we will highlight Epa9 specifically and how it can bind compared to other mutants created from the paper.
Important amino acids
The type of protein that we are looking at is an adhesion protein, so it does not function as an enzyme. It does not have a catalytic triad within the binding pocket. Though there are some important amino acid residues to highlight as they interact with the ligand (lactose). The ligand itself, lactose, is a small carbohydrate consisting of two sugars. In the diagram of the protein, we can observe the all-red ball stick structures around the ligand are the amino acid residues interacting with the ligand. These residues are They are all interacting via hydrogen bonds as depicted with this image off to the right. In the image, the hydrogen bonds involved are circled and we can see that there are only six hydrogen bonds that are binding the lactose to Epa9, which could account for a low binding affinity towards smaller carbohydrates. As it is noted that Epa9 is better at binding larger carbohydrates because of its elongated loop 1.
Structural highlights
Some things to note are that the main structure of the protein consists of . The rest of the molecule contains a primary chain structure. It can also be noted that two beta-sheets contain at least one key residue that interacts with the ligand. These parts of the beta-sheets are parts of the inner calcium-binding loops. The beta-sheets in this protein run anti-parallel and the structural shape is similar to that of a short cylinder. The structure of the stacked beta-sheets controls the shape to form binding sites. Domain A contains the binding site for carbohydrates, while the C-terminal domain contains a glycosylphosphatidylinositol (GPI) to bind the protein to the cell wall surface. The binding site in domain A has that help shape the pocket for binding the ligand. This includes two outer loops and two calcium-binding loops. The outer two loops are highly variable as compared to the highly conserved inner CBL1. The loop is conservative as it is crucial for achieving good host cell binding. The other calcium-binding loop, CBL2, is variable and tied to the protein's ligand-binding specificity. The outer pocket is made of two other loops and these are more variable, but still contain some key residues that are correlated with high binding affinity. To get a better look at the shape of the pocket there is a . We can observe that the shape of the binding pocket is dipped in and we can see where the outer two loops help shape that pocket as well as the inner CBL1 and CBL2. This allows for the space to interact with the carbohydrates on the host cell surface. Especially in Epa9 that is being represented, we can look at the elongated loop 1 in the dark navy that looks as though it cups around the pocket to interact and bind with the ligand. Because lactose is small loop 1 must curl in to help Epa9 bind to lactose. This is harder for Epa9 to do as it was found the elongated loop better for binding larger carbohydrates.
Other important features
Looking further into the structure studied we are going to compare two versions of the Epa's from the paper cited below. First looking at bound with lactose. Now looking at another structure for comparison, , but the only difference is that its CBL2 loop is from Epa1. The reason for comparing these two is to show that the change in the CBL2 being changed how by little interaction changes in the angles in the binding, that it can change the binding specificity of the protein to different carbohydrates. Remembering that this loop is a part of forming the inner binding pocket. It has a role in the binding specificity of the Epa. Exchanging of the CBL2s does not entirely change the binding specificity of the protein but makes a novel binding pattern. Comparing these two there is not much difference to the eye, but that there is one residue in CBL2 in the mixed Epa9 vs. regular Epa9. Residue 258 in the mixed Epa9 is not an asparagine, but a glutamic acid. The angle and distance that carbon 6OH in the glucose ring is from the key residues are changed partially by this, as interactions don't change but the distance in which the interaction happens will change. In the mixed Epa9, this distance change is actually why the binding pattern changes. It is to accommodate the change and find a better binding carbohydrate.
