Sandbox Reserved 1693

This is a default text for your page Sandbox 1677. Click above on edit this page to modify. Be careful with the < and > signs.

You may include any references to papers as in: the use of JSmol in Proteopedia ^[1] or to the article describing Jmol ^[2] to the rescue.

Function of your protein

The protein, L-rhamnose-a-1,4-D-glucuronate lyase, is an enzyme from the fungus Fusarium oxysporum 12S that naturally breaks down and degrades Gum Arabic (GA) by releasing the Rha caps found on the non-reducing ends of GA. I chose to specifically look at the mutant H105F Rha-GlcA ligand which has a PDB file: 7ESN. A colored image of this can be seen (For Reference N-5' = Blue, C-3' = Red)

The enzyme in this study, FoRham1 is a mutant Rha-releasing enzyme that specifically uses a-L-rhamnose-a-1,4-D-glucuronic acid (Rha-GlcA) as a .^[3] The mechanism provided in the literature shows FoRham1 removing the Rha cap from the GlcA, producing Rha sugar and GA as products. By doing this, scientist hope to be able get a better understanding of GA structure and characteristics to then advance its usefulness to society.

Biological relevance and broader implications

According to scientific literature, little is known about GA carbohydrate structure. This study is being used to understand more about the degradation/function of GA, as there is no known enzyme that can completely degrade GA. Although GA structure has been studied significantly with the use of chemical methods such as NMR and methylation, a detailed structure has not been produced as it is a complex branched polysaccharide.^[4]

Gum Arabic is produced from Acacia trees when stressful conditions (such as a drought or wound) are present.^[5] This sticky extract has many practical uses in the pharmaceutical, cosmetic, and food production industries. Serving as an effective stabilizer, emulsifier, and thickener, a better understanding of the degradation and production of GA will continue to yield beneficial industrial results.

Important amino acids

Amino acids His85, His105, Tyr150, Arg166, Ser170, Arg220, Pro223, Asn275, Arg331 are important in ligand to protein interaction. . ^[6]

His 85, Arg166, Tyr 150, and Tyr 202 are four amino acids that form hydrogen bonds with the Rha sugar. His 85 and Arg166 proved to be a key amino acid in the catalytic reaction that divides the Rha-GlcA complex in the binding pocket during the reaction. . His 105 showed to form a stabilizing hydrogen bond with His 85, reinforcing the catalytic importance of His 85. The Rha Sugar cap on the GA also interacts with amino acids Arg166, Ser170, Arg220 by Hbonding, which is an important part of separating Rha-GlcA.

Amino Acids Arg220, Pro223, Ser170, and Arg331 Hbond with the Glc independently, and they play an important role in stabilization during the catalytic mechanism. Ser170 and Pro223 Hbond with the O1 of GlcA, while Arg220 provides another hbond on the O3 atom ^[7].

Structural highlights

Secondary Structural Highlights - I found that there are lot of long antiparallel beta sheets in the protein (approx. 30 ). I also observed that there was one larger , with two smaller hydrophobic helices in the protein structure. This could mean that the protein is very stable, as antiparallel sheets provide more structure stabilization compared to parallel beta sheets. This is all due to how Hbonding between backbones occur. I also have observed that many of the beta sheets are formed of polar amino acids, which plays a role in how the protein folds.

The overall 3-D arrangement or tertiary structure of FoRam1 is quite unique. It is compose of a . This tertiary structure is unique to a bacterial family named BNR 4, and the structure is key to all of the families function^[8]. I could not find any quaternary structures for this enzyme.

The of the entire enzyme can give us a valuable insight to the structure and function relationship. As you can see from the representation, spacefill shows how deep the substrate sits in the active site. This cleft of the active is mentioned multiple times throughout the article, and the red is the Rha-Glc substrate in the active site. You can also see how the solvent (green color) interacts with the surface of the protein. These highlighted areas are often indicators of polar interactions, as the solvent (H20) is polar.

Other important features

One of the most important features of the enzyme could very easily go unnoticed. An acetate ion is pulled into the active site with the substrate Rha-GlcA, and it is a key factor in the catalytic mechanism of FoRham1. The ion interacts with the GlcA and acts as a neutralizer during the catalytic reaction that separates Rha-GlcA. I could not create a visualization of this as the PDB file places the acetate ion outside of the active site.

Another very interesting and important feature was the active site location. You can find the active site of the enzyme in the cleft of the center on one of B-propeller blades. The depth and location of the cleft/active site is important for substrate binding and the catalytic reaction, as the shape is determined by the amino acids. This shape then determines the function, thus showing how important the cleft is to the enzyme.