BENG 230A – Biochemistry Midterm Review Fall 2022
Biomolecules & Cell Chemistry
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What is in a cell?
Glycerol + Fatty Acids + Phosphate group
Plasma Membrane, Signaling Molecules, Hormones
Sugar (Ribose/Deoxyribose )
+ Phosphate group + Nitrogenous base (A/T/G/C/U)
Material (DNA and RNA)
Polymers of monosaccharides linked by glycosidic bonds
Energy production and storage (glycolysis), Post translational modifications (glycosylation)
Polymers of
amino acids linked by peptide bonds
Most abundant molecule (70% of cell mass)
Universal solvent, medium of transportation between intracellular and extracellular compartments
Enzymes, Antibodies, Hormones, Cytoskeleton
Covalent and non-covalent interactions in biological systems
Hydrogen bonding
Difference in electronegativity
Van der Waals Interactions
Distance dependent interaction of oppositely polarized electron clouds
Hydrophobic Interactions
Polar groups – Hydrophilic Non-polar groups – Hydrophobic
Strength: Covalent > Ionic > Hydrogen Bond > Hydrophobic > Van der Waals
Protein Structure & Function
Amino Acids Classification
● Amino acids + Interactions: Determine protein structure
● Changes in amino acid sequence -> Misfolding of protein -> Loss of function
3D Structure of Proteins
Energetically favorable conformation
Central Dogma of Molecular Biology
From DNA to DNA: Replication
DNA replication Separation, Base pair
DNA Synthesis by DNA polymerase
DNA replication Fork
DNA Proofreading
Why 5’->3’?
The need for accuracy
DNA Replication at the Lagging strand
Done by DNA Ligase
DNA Helicase
DNA double helix are tightly coupled. High temperature is needed to break them (95oC)
DNA Binding Protein
SSB: Single Strand DNA-binding Proteins, also called helix destabilizing proteins
DNA Clamping Protein
Protein machinery for DNA replication
Telomerase and its function
Retrovirus-based Transposition
Non-retroviral retrotransposition
From DNA to RNA: Transcription
DNA->RNA-> Proteins
Genes expressed with different efficiency
The chemical structure differences between
DNAs and RNAs 1. ribose, deoxyribose 2. Uracil and thymine
RNA base pairs A-U; G-C
RNA Structures
Initiation of transcription with RNA polymerase II in eucaryotes
TF: transcription factor TBP: TATA box binding protein Promoter upstream of real starting sequence of transcription
TFIIH open DNA double helix and phosphorylate C-tail of polymerase and allow the release and transcription
The importance of RNA polymerase II tail
Initiation of transcription with RNA polymerase II in eucaryotic cells Remember Nucleasomes
Enhancer, mediator, chromatin remodeling complex, histone acetylase
mRNA between procaryotic and eucaryotic cells 5’ capping and 3’ polyadenylation
Genes to proteins
The comparison between eucaryotes (substantially complex) and procaryotes (simple)
From RNA to Protein: Translation
Key Player #1: Transfer RNA (tRNA)
Amino acid attachment and interaction with the ribosome
Interacts with mRNA
Aminoacylation of tRNA by aminoacyl-tRNA synthetase
Proofreading ability for translational fidelity
Key Player #2: Messenger RNA (mRNA)
The Genetic Code
Wobble position in codon
20 Amino Acids, 64 Codons Redundancy but no ambiguity
Key Player #3: Ribosome
Protein synthesizing organelle
Aminoacyl site Peptidyl site Exit site
Ribosomal RNA (rRNA) – Binds to tRNA and mRNA to ensure accurate translation
Translation: Initiation, elongation, release
Start codon: AUG, codes for transferase activity of large ribosomal subunit (ribozyme)
Ribosomal translocation
Stop Codons: UAA, UAG, UGA
From DNA to Protein: Techniques
Why do cells in your body behave differently despite having mostly identical genome?
Evaluation: An Overview
For each of the key technique, you need to master:
• Use case
• Pitfalls
• Compensation methods
A good format to following when describing an experiment on the test:
• Control/experimental groups
• Technique
• Expectation from analysis
• pitfalls
RT-qPCR: An Overview
RT-qPCR: Things to Consider
• Limitations in target selection (primers) • Scalability
• Normalization across samples/genes
Bulk v.s. Single-Cell RNA Sequencing
Sample scRNA-seq Output
Nona Farbehi, , , , , -Lis, K Ho, Nordon, Harvey (2019) Single-cell expression profiling reveals dynamic flux of cardiac stromal, vascular and immune cells in health and injury eLife 8:e43882
https://doi.org/10.7554/eLife.43882
Sample scRNA-seq Output
Nona Farbehi, , , , , -Lis, K Ho, Nordon, Harvey (2019) Single-cell expression profiling reveals dynamic flux of cardiac stromal, vascular and immune cells in health and injury eLife 8:e43882
https://doi.org/10.7554/eLife.43882
Bulk RNA-seq: Things to Consider
Sequencing depth
Read length
– Gene1: ACAAA Gene2: GGAAA – Read1: AAA Read2: CAAA
• Heterogeneity
Normalization across samples
– Gene1: 100/1000 reads Gene2: 200/2000
scRNA-seq: Things to Consider
Extreme cost Dropout effect Batch effect
Complete list of challenges can be found here:
Eleven grand challenges in single-cell data science | Genome Biology | Full Text (biomedcentral.com)
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