Lecture 4_BIOC212_Maria Vera Ugalde_2025 - Copy
Page 1
Title
Protein Folding in the Cell - 2
Course: BIOC212 Winter 2025
Instructor: Maria Vera Ugalde
Page 2
Summary from Lecture 3
Amino acids (AA) bind through peptide bonds; sequence determines protein folding into functional 3-D structures.
Both AA peptide bonds and side chains engage in non-covalent interactions to form secondary, tertiary, and quaternary protein structures.
The side chain (R) influences protein charge, polarity, and hydrophobicity, affecting interactions.
Similar AAs provide homology among protein family members or conserved proteins through evolution:
Types of AAs:
Hydrophobic
Polar
Basic
Acidic
Proteins are polymers made of 20 different amino acids, categorized by functional group of side chains.
Page 3
Sequence Similarity
Sequences of polypeptides can be compared to align identical and similar amino acids.
Sequence similarity (homology) indicates evolutionary conservation.
Homology suggests common structure or function.
Polypeptides without sequence similarity are divergent.
Page 4
Example of Sequence Similarity
Example sequences from Homo sapiens and Apis mellifera:
Alignment reveals identity, strong similarity, and weak similarity.
Page 5
Amino Acid Side Chains
Uncharged Polar Side Chains:
Asparagine (Asn, N), Glutamine (Gln, Q)
Nonpolar Side Chains:
Valine (Val, V), Alanine (Ala, A), Leucine (Leu, L), Glycine (Gly, G), Cysteine (Cys, C), Isoleucine (Ile, I)
Basic Side Chains:
Lysine (Lys, K), Arginine (Arg, R), Histidine (His, H)
Acidic Side Chains:
Aspartic acid (Asp, D), Glutamic acid (Glu, E), Methionine (Met, M)
Page 6
Protein Families
A family consists of proteins or domains with homologous sequences and structures, often with related functions.
An organism may have multiple proteins from the same family, found across different organisms.
Page 7
Topics for Discussion
Post translational modifications of AAs.
Protein folding processes.
Protein misfolding.
Protein quality control mechanisms.
Assisted folding by chaperones.
Page 8
Post-Translational Modification (PTM)
Proteins can undergo chemical modification post-translation, contributing to diversity and regulating function.
Types of modifications include:
Cleavage into smaller proteins by peptidases.
Covalent modifications of N-terminus or side chains.
Page 9
Functions of Side Chain Modifications
Can alter protein surface or conformation, create/block binding sites, and are often regulated and reversible.
Modifications are swift, acting as functional switches.
Main types: Phosphorylation, Methylation, Acetylation, Glycosylation, Sumoylation, Ubiquitination.
Page 10
Phosphorylation
Major regulatory mechanism involving phosphorylation of hydroxyl groups (S, T, Y).
Kinases transfer phosphates from ATP and are specific to side chains and surrounding sequences.
Phosphatases remove phosphate groups.
Page 11
Phosphorylation Enzymes
Kinase Families:
Ser/Thr kinases, Tyr kinases, dual specificity kinases.
Phosphatase Families:
Ser/Thr phosphatases, protein Tyr phosphatases, dual specificity phosphatases.
Study of phosphorylation's role in protein function; phosphomimic and de-phosphorylated strategies.
Page 12
Phosphopeptide Binding
Specialized domains bind phosphorylated serine, threonine, or tyrosine.
Phosphorylation is vital for binding; surrounding polypeptide sequences also contribute.
Page 13
Acetylation of Lysine
Acetylation changes polarity and involves lysine acetyltransferases (KATs) and deacetylases (KDACs).
Effects include increased size and charge, altering function such as in histone acetylation.
Page 14
Methylation of Lysine & Arginine
Methylation adds 1-2 methyl groups, creating methylarginines.
Lysine can be mono-, di-, or trimethylated; involves specific methyltransferases and demethylases.
Page 15
PTM and Binding
Acetylation and methylation provide new binding sites; specific domains must bind modified residues and surrounding sequences.
Page 16
Protein Folding and Structure
Folded protein structure derives from hydrophobic interactions internally, with polar side chains on the exterior.
The native state represents the fully folded conformation of a protein, where modifications may occur post-folding.
Page 17
Importance of Interactions for Folding
Hydrogen bonds stabilize secondary structures and involve peptide bonds.
Hydrophobic interactions dictate tertiary structures and involve side chains.
Various bonds (hydrogen, ionic, disulfide) contribute to protein stability.
Page 18
Relationship Between AA Sequence and 3D Structure
Native structure determined by primary AA sequence; represents a state of minimal energy and is thermodynamically favored.
Although folding can be spontaneous, biological processes assist in folding.
Page 19
The Folding Process
Folding is complex and transitions through intermediates with increasing structure toward the native state.
Unfolded domains show extended conformations lacking secondary or tertiary structure.
Page 21
Native State of Proteins
The native state is the most stable conformation stabilized by hydrophobic contacts and sometimes requires a ligand partner.
Equilibrium can exist between native and near-native states.
Page 22
Folding Intermediates
Intermediates have some secondary structure but incomplete tertiary structure, exposing hydrophobic regions.
Increased flexibility leads to aggregation risks and insolubility due to hydrophobic contact preferences.
Page 23
Protein Misfolding Issues
Causes of misfolding include incomplete folding, unavailable ligands, or genetic mutations.
Misfolded proteins may lead to diseases and are exacerbated by environmental factors, aging, and decreased protein quality control efficiency.
Page 24
Genetic Mutations
Mutations alter polypeptide sequences (substitution, insertion, deletion) and affect protein folding or function.
Some mutations are pathogenic, while others may not show significant effects.
Page 25
Protein Production Rates
Nucleus: ~1000 transcripts/second.
Total mRNAs and proteins reach 10^5-10^6 and 10^10, respectively.
Page 26
Protein Quality Control and Homeostasis
Chaperones are central to maintaining protein quality, assisted by proteasomes and autophagy.
Proteostasis ensures correct protein concentration, conformation, and location, vital for proteome stability.
Page 27
Chaperones and Their Role
Molecular chaperones aid in folding and prevent aggregation without integrating into the native state; often recognize exposed hydrophobic regions.
Heat shock proteins (HSP) are key chaperones expressed during stress conditions.
Page 28
Stress Response Mechanisms
Cells respond to stress via increased expression of chaperones, forming the Heat Shock Response (HSR) and Unfolded Protein Response (UPR).
HSR protects against cell death; UPR manages ER protein stress.
Page 29
Activation of Chaperones
Inducible chaperones are activated by heat and stress; constitutive chaperones assist general protein folding under non-stress conditions.
Page 30
Unfolded Protein Activation
Unfolded cytosolic proteins activate heat shock responses and transcription of HSPs, adjusting protein synthesis based on stress severity.
Page 31
HSF1 and Heat Shock Response
HSF1 mediates heat shock response and is regulated through interactions with Hsp90, influencing its activation and down-regulation mechanisms.
Page 32
Regulation of HSF
Active HSF1 trimerizes for transcription activation after heat shock, whereas monomers bind Hsp90 to maintain its inactive state.
Page 33
Phosphate Binding Amino Acids
Identifying amino acids that bind phosphate groups, specifically phospho-tyrosine.
Page 34
Thought Questions
Identify amino acids involved in binding to phospho-serine.
Compare recognition between phospho-serine and phospho-tyrosine.