Episode 6: Into the Darkness

The meme of the week is definitely gonna be grinds my gears. A classic from family guy where people express their discontent.

Subtopic 1: Protein engineering and folding history.

Start with this because it sets up the background and creates the significance for my research. This topic will explain the goals of protein folding and how it has progressed through the years.

1. Protein engineering’s end goal is to make new proteins or new functions out of existing proteins or from scratch
1. Directed Evolution is a way to achieve that goal
1. Lutz discusses the process of directed evolution and the history of directed protein evolution. They talk about the computational tools for the evaluation of protein data sets. The methodological advances in semi-rational enzyme engineering and de novo enzyme design in recent years provide researchers with powerful and effective new strategies to manipulate biocatalysts. Directed evolution represents an obvious strategy to improve existing designer enzymes and to potentially “break” the performance cap.
2. Packer talks about pioneering studies in the field of directed evolution sought to improve the wild-type activity of enzymes through the enhancement of solubility, thermostability, affinity for substrate or catalytic turnover in simulations. The future of new directed evolution studies is bright.
2. In order to create new proteins, we must correctly predict the structure of the protein, which is no easy task.
2. 60 years ago, there were no discernable methods to figure out the specifics of protein folding, through Moore’s Law the methods will be more advanced in the future
1. Dill talks about Moore’s Law and the benefits of folding in the future, including the possibility of solving folding failures like Parkinsons and Alzheimers
3. Currently, there are many methods but experimentation and computer modeling are leading the way.

Subtopic 2: Different prediction methods Computer Simulation vs Experimentation.

1. Experimentation in labs is more Accurate
1. This only results in a small amount of structures because it is time consuming (2500)
1. Kumar
2. Computer analysis methods are less accurate now, but have the potential to create much more structures in a much faster manner in the future
1. They are also currently the most accessible form of analysis
1. Need to back this up
3. Thus computer models are the future of this field
1. CASP competition
1. Recently the computer based prediction models are more accurate than other methods
1. Kumar and CASP
2. Because computer based models are the more effective long term solution, I will be using them
2. REMD, SIFT, FIRST
1. I need to look into this some more and compare these methods, I just found some articles for this
2. FIRST, and REMD
1. De Graff
1. Crack Propagation
2. Thorpe
1. New methods for protein folding by using short-range forces in proteins modeled by constraints. Forces included in the analysis are the covalent bond-stretching and bond-bending forces, salt bridges, and hydrogen bonds. They used colored mapping to provide a visualization of the motion of the protein.
2. FIRST stands for Floppy Inclusion and Rigid Substructure Topography.

Subtopic 3: LAC1 Repressor, why look at it?

1. Definition of LAC1 Repressor
1. The crystal structure of the lac repressor is a bent structure with all 4 of the DNA binding portions pointing in one direction.
2. The DNA sequence has 3 lac repressor recognition sites within 500 base pairs. In the quaternary formation of the repression loops, the repressor interacts simultaneously with 2 genomic DNA sites.
3. Lewis
2. The repressor is important because it suppresses the coding for proteins that break down glucose, galactose, and other sugars.
1. Prevents the creation of Beta-galactosidase, which is the enzyme that performs the first step in lactose metabolism, making glucose and galactose. Galactoside acetyltransferase, which actually doesn't really do much in lactose metabolism (its unclear). And lactose permease transports lactose through membranes.
2. LAC1 plays a key role in lactose breakdown, understanding exactly why the protein functions the way it does will provide insight into diseases such as Lactose intolerance.
3. Goodsell from the Protein Database
4. Possibly engineer the LAC1 into not repressing those enzymes

Subtopic 4: Conserved vs Nonconserved positions

1. Certain parts of protein are essential to function, others are not
1. There is a dispute on whether or not all segments of the protein are essential to function
2. Conserved positions control major folding points that change the entire structure. Nonconserved positions control less important points in the protein that do not contribute to major structural problems.
1. However the function may be affected by Nonconserved position
3. Over many years scientists have attempted to change nonconserved positions to fine tune function.

Subtopic 5: The significance of Structure for Function

1. The structure of the protein is essential for understanding the biological role of the protein. Structural genomics aims to structurally characterize most protein sequences by a combination of experimentation and prediction
1. Baker
2. Could be placed after Subtopic 1

Combining both the significance of protein folding and prediction with the protein of LAC1 leads us to investigate the importance of Conserved vs nonconserved positions. Thus the question: Do nonconserved positions, when mutated, affect the function of the protein more significantly than conserved positions in LAC1.

Purpose: Understand definitions and the methods for testing LAC1. And understanding the future implications of the results whether it corroborates common evidence or not.

So I got a lot of subtopics, but I feel like the order could be moved around, if you could help me out that would be great. Help me out friends.