Week 4 HW: Protein Design Part I

Part A. Conceptual Questions Answer any NINE of the following questions from Shuguang Zhang: (i.e. you can select two to skip)

Why do humans eat beef but do not become a cow, eat fish but do not become fish? Even though some may like to say you are what you eat, this really isn’t the case when it comes to eating an organism. Even though we consume its parts and convert protein into energy we are transforming energy into energy that we can use. One may argue we embody that organism to a degree, but we dont fully transform into that being. Your cells follow a genetic blueprint, and while other DNA enters your body, you do not absorb its genome, nor does it integrate into yours.

Why are there only 20 natural amino acids? Although the genetic code contains 64 possible codons, evolution stabilized around 20 canonical amino acids. This was not due to a strict numerical limitation, but because these 20 provide an optimal balance of chemical diversity, stability, and functional efficiency. Biochemical availability on early Earth also played a crucial role, early life utilized the amino acids that were most readily synthesized under prebiotic conditions. Mathematical analyses of the canonical set suggest that the 20 amino acids achieve a near-optimal coverage of chemical property space, maximizing structural and catalytic versatility while minimizing redundancy. Furthermore, the structure of the genetic code itself is organized in a way that reduces the impact of mutations: codons that differ by a single nucleotide often encode amino acids with similar physicochemical properties, thereby limiting potential damage to protein structure and function.

Can you make other non-natural amino acids? Design some new amino acids. Yes, and we already have. Scientists have even been able to genetically engineer organisms to expand the genetic code beyond the 20, by introducing new codons.() Some of the amino acids that have been synthesised include fluorescent amino acids, for example, to help scientists localise proteins in a cell and observe their interactions, improving observation.

Where did amino acids come from before enzymes that make them, and before life started? Amino acids can form spontaneously under prebiotic conditions through chemistry. Scientists have simulated Earth’s atmosphere using simple gases and found that amino acids can form through life’s basic chemistry.

If you make an α-helix using D-amino acids, what handedness (right or left) would you expect? If you make an α-helix using D-amino acids, you would expect a left-handed twist, because the molecular building blocks themselves are mirrored. D-amino acids have the opposite chirality of the natural L-forms, so their backbone geometry favors a left-handed helix rather than a right-handed one.

Can you discover additional helices in proteins? Yes research continues to identify unsual or distrorted helical geometries. Scienttists have also designe helices from non antural amino acids, or even alternate hydrogen bonding patterns.

Why are most molecular helices right-handed? Most moleculart helices are right handed beacuse of chirality. The presence of molecular assymetry in L- amino acids determening helix twist direction, favouring a right handed direction. While left handed helices are possible, they are rare. An example of a left handed helices is a D helices, which its molecular building block is most stable and corresponds to a left hand.

Why do β-sheets tend to aggregate? β-sheets tend to aggregate because their extended structure exposes backbone hydrogen bond donors and acceptors, allowing strands from different proteins to easily form intermolecular hydrogen bonds. Their flat geometry also enables tight stacking into stable, repetitive “cross-β” structures, making aggregation energetically favorable.

What is the driving force for β-sheet aggregation? Backbone hydrogen bonds form between neighbouring β strands, satisfying exposed N-H and C=O groups. Hydrophobic chains pack together and repel water, thus loweing the systems free energy and agregates state is more stable than partially unfolded state.

Why do many amyloid diseases form β-sheets? Because the β-sheet structure is the most stable form in which misfolded proteins can agregate and be reorganized, creating a strong cross β structure.

Can you use amyloid β-sheets as materials? Amyloid β-sheets are one of the strongest and most stable protein structures in nature, making them greatly appealing in material architecture. Some of the properties that arise and be applied in material science include, tensile strength, chemical stability, self assembly in asolutions, heat resistance, and mechanical stiffness. Examples where nature uses the intelligent structure present in β-sheet scaffolidng is in silk fibroin and curli fibers in bacterial biofilms. Common applications of Amyloid β-sheets are hydrogels, drug delivery, nanofibres, tissue scafolidng, bioprinting, and even in conductive biofilms.

Design a β-sheet motif that forms a well-ordered structure.

Part B: Protein Analysis and Visualization

1. Briefly describe the protein you selected and why you selected it.

2. Identify the amino acid sequence of your protein. 2.1 How long is it? What is the most frequent amino acid? You can use this Colab notebook to count the frequency of amino acids. 2.2 How many protein sequence homologs are there for your protein? Hint: Use Uniprot’s BLAST tool to search for homologs. 2.3 Does your protein belong to any protein family?

3. Identify the structure page of your protein in RCSB 3.1 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) 3.2 Are there any other molecules in the solved structure apart from protein? 3.3 Does your protein belong to any structure classification family?

4. Open the structure of your protein in any 3D molecule visualization software: 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)?