Week 4 HW: Protein Design Part I

Homework Questions

Part A. Conceptual Questions

  • 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.

  • 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. 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 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 Does your protein belong to any protein family? Identify the structure page of your protein in RCSB 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) Are there any other molecules in the solved structure apart from protein? Does your protein belong to any structure classification family? 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”. Color the protein by secondary structure. Does it have more helices or sheets? Color the protein by residue type. What can you tell about the distribution of hydrophobic vs hydrophilic residues? Visualize the surface of the protein. Does it have any “holes” (aka binding pockets)?

Part C. Using ML-Based Protein Design Tools

HTGAA ProteinDesign2025

Fold your protein with AlphaFold or ESMFold or Boltz and compare it to the real structure. Comment on: Any predicted vs. experimental differences. Low-confidence regions and why do you think they are low confidence? Inverse-fold your structure with ProteinMPNN What sequence do you get? Is it the same as the original sequence you folded? Why yes or no?

Part D. Group Brainstorm on Bacteriophage Engineering

Find a group of ~3–4 students Review the Bacteriophage Final Project Goals: Increased stability (easiest) Higher titers (medium) Higher toxicity of lysis protein (hard) Brainstorm Session Choose one or two main goals from the list that you think you can address computationally (e.g., “We’ll try to stabilize the lysis protein,” or “We’ll attempt to disrupt its interaction with E. coli DnaJ.”). Write a 1-page proposal (bullet points or short paragraphs) describing: Which tools/approaches from recitation you propose using (e.g., “Use Protein Language Models to do in silico mutagenesis, then AlphaFold-Multimer to check complexes.”). Why you think those tools might help solve your chosen sub-problem. One or two potential pitfalls (e.g., “We lack enough training data on phage–bacteria interactions.”). Include a schematic of your pipeline This resource may be useful: HTGAA Protein Engineering Tools Individually put your plan on your website page Each group’s short plan for engineering a bacteriophage Schedule time ( HTGAA Protein Engineering Feedback) to get feedback/discuss your ideas, and put the feedback on your website