Homework Week 4 part A,B,C

Part A. Conceptual Questions

Answer any of the following questions by Shuguang Zhang:
- How many molecules of amino acids do you take with a piece of 500 grams of meat? (on average an amino acid is ~100 Daltons)
- Why humans eat beef but do not become a cow, eat fish but do not become fish?
- Why there are only 20 natural amino acids?
- Can you make other non-natural amino acids? Design some new amino acids.
Yes, it is possible to design and synthesize non-natural amino acids (NNAAs), also known as unnatural or non-canonical amino acids. These amino acids are not among the 20 standard ones encoded by the universal genetic code but can be engineered to introduce novel functionalities into proteins.

The synthesis of peptides and proteins containing non-natural amino acids

Reprogramming natural proteins using unnatural amino acids

Stereoselective amino acid synthesis by synergistic photoredox-pyridoxal radical biocatalysis
- Where did amino acids come from before enzymes that make them, and before life started?
- If you make an alpha-helix using D-amino acids, what handedness (right or left) would you expect?
- Can you discover additional helices in proteins?
- Why most molecular helices are right-handed?
- Why do beta-sheets tend to aggregate?
  - What is the driving force for b-sheet aggregation?
- Why many amyloid diseases form b-sheet?
  - Can you use amyloid b-sheets as materials?
- Design a b-sheet motif that forms a well-ordered structure.

Part B: Protein Analysis and Visualization

In this part of the homework, you will be using online resources and 3D visualization software to answer questions about proteins.

Pick any protein (from any organism) of your interest that has a 3D structure and answer the following questions.
- Briefly describe the protein you selected and why you selected it.
I have selected the cutinase enzyme from Thermobifida fusca for discussion because its notable role in biodegrading synthetic polyesters, particularly polyethylene terephthalate (PET). This enzyme catalyzes the hydrolysis of cutin, a structural polymer in plant cuticles, and has been found effective in breaking down PET, a common plastic. Its potential applications in plastic waste management and environmental remediation make it a significant subject of study.

Recent trends in microbial and enzymatic plastic degradation: a solution for plastic pollution predicaments - Biotechnology for Sustainable Materials

Engineering Plastic Eating Enzymes Using Structural Biology

Plastic-Degrading Enzymes from Marine Microorganisms and Their Potential Value in Recycling Technologies
- Identify the amino acid sequence of your protein.
  - How long is it? What is the most frequent amino acid? You can use this notebook to count most frequent amino acid - https://colab.research.google.com/drive/1vlAU_Y84lb04e4Nnaf1axU8nQA6_QBP1?usp=sharing
  Long: 261 amino acids. The most frequent amino acidis Serine (S), appearing 26 times.
  - How many protein sequence homologs are there for your protein?Hint: Use the pBLAST tool to search for homologs and ClustalOmega to align and visualize them. Tutorial Here
  At least 22 with 95% of percentage of identity.
  - Does your protein belong to any protein family?
  It belongs to the cutinase family, which is a subset of the larger α/β-hydrolase superfamily. Cutinases are serine esterases characterized by a catalytic triad composed of serine, histidine, and aspartic acid residues. They are primarily known for hydrolyzing cutin, a structural polyester in plant cuticles, but also exhibit activity toward various esters and synthetic polyesters.
  
  Cutin is a waxy, water-repellent biopolymer found in the cuticles of plants, covering their aerial surfaces such as leaves, stems, and fruits. It serves as a protective barrier against environmental factors, including water loss and microbial invasion.
  
  Frontiers | The Plant Cuticle: An Ancient Guardian Barrier Set Against Long-Standing Rivals
- Identify the structure page of your protein in RCSB
https://www.rcsb.org/structure/5ZOA
- When was the structure solved? Is it a good quality structure? Good quality structure is the one with good resolution. Smaller the better (Resolution: 2.70 Å)
Deposited: 2018-04-12 Released: 2019-04-17
- Are there any other molecules in the solved structure apart from protein?
Yes, apart from the protein itself, the solved structure of 5ZOA includes chloride ions (Cl⁻) as non-protein molecules. These chloride ions may play a role in stabilizing the structure or influencing enzymatic function. However, the structure does not contain additional ligands, cofactors, or small molecules like substrates, inhibitors, or metal ions.
- Does your protein belong to any structure classification family?
It belongs to the α/β-hydrolase fold superfamily.
- Open the structure of your protein in any 3D molecule visualization software:
  - PyMol Tutorial Here (hint: ChatGPT is good at PyMol commands)
  - Visualize the protein as "cartoon", "ribbon" and "ball and stick".
  Cartoon
  
  Ribbon
  
  Ball and stick
  - Color the protein by secondary structure. Does it have more helices or sheets?
  Red represents α-helices, yellow represents β-sheets, and green represents loops/coils. The structure has more α-helices (red) than β-sheets (yellow)
  - Color the protein by residue type. What can you tell about the distribution of hydrophobic vs hydrophilic residues?
  - White → Hydrophobic (nonpolar) residues (Ala, Val, Leu, Ile, Pro, Phe, Met, Trp)
  - Blue → Negatively charged (acidic) residues (Asp, Glu)
  - Red → Positively charged (basic) residues (Lys, Arg, His)
  - Green → Polar (hydrophilic) residues (Ser, Thr, Tyr, Asn, Gln, Cys)
  Hydrophobic Residues (White) → 58.94%
  
  Hydrophilic Residues (Green) → 41.06%
  
  Plastics like polyethylene (PE), polypropylene (PP), and PET (polyethylene terephthalate) are highly hydrophobic materials. For a protein (enzyme) to degrade plastic efficiently, it needs to interact with the plastic surface. This interaction is stronger if the enzyme has a significant hydrophobic surface.
  
  58.94% hydrophobic residues → Good for interacting with plastic surfaces
  
  41.06% hydrophylic residues → Potentially useful for maintaining solubility in water
  
  If the active site or surface of your protein has many hydrophobic residues, it can bind to plastics more effectively. If some polar residues are near the active site, they may help catalyze hydrolysis or oxidation reactions, which are important for breaking down plastic.
  
  Are the hydrophobic residues on the surface or buried inside the protein?