BIOSC 1540: L11 (Structural biology)
This is a live streamed presentation. You will automatically follow the presenter and see the slide they're currently on.
This is a live streamed presentation. You will automatically follow the presenter and see the slide they're currently on.
Computational Biology
(BIOSC 1540)
Oct 8, 2024
Lecture 11:
Structural biology
At the foundation of biological processes lie atoms and their interactions
Why is Green Fluorescent Protein (GFP) fluorescent, but not the chromophore in solution?
GFP keeps the chromophore planar and facilitates an excited-state proton transfer
Fluorescent
Not Fluorescent
Covalent bonds are formed when atoms share pairs of electrons that holds molecules together
Strength and stability: Covalent bonds provide the necessary stability for complex biological structures
Directionality: Covalent bonds limit the specific angles and orientations leading to the 3D shapes of biomolecules
Noncovalent interactions are weaker than covalent bonds and involve electrostatics
We will cover this in a later lecture
Macromolecular structure
Membrane Formation
Protein-Protein Interactions
Molecular recognition
The precise arrangement of atoms determines how molecules fold, bind, and perform biological tasks
CRISPR-Cas9
COVID-19 treatments
High-throughput sequencing
Innovation and biotechnology depend on molecular understanding
Structural biology determines the 3D shapes of biological macromolecules and how these shapes relate to function
Why study structure?
Primary Goal: To understand how molecular machines in cells work by deciphering their atomic arrangements.
The primary structure of a protein is the linear sequence of amino acids, held together by covalent peptide bonds
The primary structure is crucial because it dictates how the protein will fold into higher-order structures
The primary structure alone does not reveal the protein's functional form or activity
While the primary sequence is critical, the folding process may also depend on cellular factors (e.g., chaperones)
The secondary structure refers to local conformations of the polypeptide chain, stabilized primarily by hydrogen bonds
Secondary structures can undergo local fluctuations—alpha helices can unwind, and beta-sheets can twist—adding to functional flexibility
These structural motifs are critical for certain functions
While alpha-helices and beta-sheets dominate, other structural motifs (e.g., 310 helices) are less common and sometimes overlooked
The tertiary structure refers to the complete 3D shape of a single polypeptide chain
Tertiary structures reveal active sites or binding pockets where catalysis or molecular interactions occur
Predicting how a sequence folds into its tertiary structure is complex, even with knowledge of secondary structures
Principle quantum number
1
2
3
Orbital quantum number
0
-1
0
1
-1
0
1
-2
2
Magnetic quantum number
1
1
2
3
2
1
(You don't need to know what these mean)
An electron at (n, l, m) will have a specific energy level and characteristics
Benzene has . . .
Six carbon atoms with 1s2 2s2 2p2
Six hydrogen atoms with 1s
located at the center of each atom's position
Particles (e.g., electrons and photons) can interact with these molecular orbials
D6h structure
D3h structure
These molecular orbitals determine behavior
Changing the positions (or symmetry) change molecular orbitals
All experimental techniques are based on probes interacting with molecule's electron density to reveal structural information
Electron density of benzene
Basic Principle: Photons scatter when they interact with atoms
The scattered X-rays form a diffraction pattern unique to the crystal
Probe: Photon (carrier of electromagnetic radiation)
What happens when two waves overlap?
If wavelengths are similar and in phase, they constructively interfere
If waves are out of phase, they deconstructively interfere
If wavelengths are similar and in phase, they constructively interfere and form spots based on atom type and distance
The spots on the detector represent the reflections of the scattered X-rays
The diffraction pattern does not directly show the atomic positions, but provides the data needed to infer the electron density
The 3D electron density map reveals the distribution of electrons in the crystal, indicating where atoms are located
The electron density map is interpreted by fitting atomic models (e.g., amino acids for proteins) into the density
Low-resolution data make it difficult to assign atomic positions precisely, leading to uncertainty in the model
Molecules
Crystals
Crystals have the same repeating unit cell, which amplifies our signals
If in solution, particles would be
Let's find information about our project's drug target: Dihydrofolate reductase
UniProt is a comprehensive database to access curated data about protein structures, functions, sequences, and annotations.
This page shows the results of a search in UniProtKB for a specific protein, in this case, "Dihydrofolate reductase"
On the left side, you have multiple filters to narrow your search results:
Reviewed (Swiss-Prot): Experts manually curated and verified these entries, ensuring high accuracy
Unreviewed (TrEMBL): These entries are automatically generated and have not been manually reviewed
Each row in the table represents a different protein entry
Entry ID: A unique identifier for the protein (e.g., P00383). You can click on this ID for detailed information about the protein
In Cryo-EM, a beam of high-energy electrons is used instead of photons
Why Electrons?
No crystals: The sample is rapidly frozen in vitreous ice to preserve its native structure
Single Particle Analysis is the main Cryo-EM technique used to determine the 3D structures of individual macromolecules
Molecules are not static
Example: The p53 tumor suppressor protein has flexible regions critical for its regulation and binding interactions
Proteins often exhibit flexibility, disordered regions, and multiple conformations
Why It Matters: Structural techniques often require ordered or stable configurations
One strength of Cryo-EM is its ability to capture multiple conformational states of a molecule, providing insights into flexibility and structural heterogeneity.
Challenge: A major issue in Cryo-EM is that highly flexible or disordered molecules may appear as fuzzy or low-resolution regions in the final structure
Advanced computational techniques are required to sort out different conformations present in the Cryo-EM data
Intrinsically disordered proteins (IDPs) or regions lack a stable 3D structure under physiological conditions but are still functional, often gaining structure upon binding to partners
Many proteins function by switching between different conformations, which is essential for their activity (e.g., enzymes, transporters, and receptors).
Technical Limitations
Biological Complexity
Resource Constraints
Lecture 11:
Structural biology
Today
Thursday
Lecture 12:
Protein structure prediction
