Note For Biochemistry

This post is writing during the teaching of the professors from The University of Manchester.

Maybe it is really puzzling as which is a combination of Structure Biochemistry, Cell Biology and Molecular Biology.

A Tour of the Cell

Need to re-write according the record.

Cytoskeleton

Components of the cytoskeleton

1. Microtubules
   Made of tubulin, 25nm diameter
   e.g. mitosis, cilia, flagella, centrioles
2. Actin filaments (Microfilaments)
   Made of actin, 6nm diameter
   e.g. cytoplasmic streaming
3. Intermediate filaments
   Several proteins such as keratin
   Intermediate in size i.e. 10 – 15nm diameter
   e.g. Keratins in skin, hair, nails, (animal horn) etc

Membrane Structure and Function

Dr. Katie Finegan

Intended Learning Outcomes

Students should be able to:

1. Describe the function and components of the plasma membrane
2. Explain how endocytosis and exocytosis facilitate the movement of large biomolecules and micro-organisms across the plasma membrane
3. Explain what is meant by: simple diffusion, facilitated diffusion and active transport
4. Differentiate between carrier proteins & channel proteins
5. Understand how membrane transporters can create and maintain solute gradients e.g. Na^+^/K^+^ pump
6. Give an example of a drug which exerts its effect by blocking membrane transport
7. Give an example of a disease where MDR transporters cause drug resistance to develop

Cell Membrane / Plasma Membrane

1. Function: Regulates materials entering and exiting the cell.
2. Structure: Two layers of phospholipids, proteins

DNA Composition and Structure

Dr. Katie Finegan

Intended Learning Outcomes

Describe the structure of DNA
Describe how DNA replicates
Describes how DNA fits into chromosomes

Nucleotides

The phosphate and sugar form the backbone of the DNA molecule, whereas the bases form the “rungs”.

There are four types of nitrogenous bases.

Each base will only bond with one other specific base.

Adenine (A)
Thymine (T)

Cytosine (C)
Guanine (G)

DNA Structure

DNA consists of two molecules that are arranged into a ladder-like structure called a Double Helix.

A molecule of DNA is made up of millions of tiny subunits called Nucleotides.

Each nucleotide consists of:

• Phosphate group
• Pentose sugar
• Nitrogenous base

Because of this complementary base pairing, the order of the bases in one strand determines the order of the bases in the other strand.

The strands are held together by complementary base-pairing

AT base pairs are held together by 2 hydrogen bonds

GC base pairs are more strongly bonded with 3 hydrogen bonds

This was consistent with the earlier observation by Chargaff that in all DNA:

%A = %T and %G = %C

Chargaff’s Rule

—**% of Guanine and Cytosine are equal**

—**% of Adenine and Thymine are equal**

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Post-Translational Modification

Genome → Transcriptome → Proteome

In the last decade, it has been found that the human proteome is much more complex than the human genome.

A single gene can give rise to multiple RNA transcripts…

each transcript can be modified to give multiple versions of a single protein

Post-translational Modification (PTM) is of fundamental importance for biological activity of proteins

PTM plays a key role in the activity, localization & interaction of proteins with other cellular molecules

Modifications have significant effects on protein structure and function and are of concern for proteins of therapeutic or diagnostic use

The proteome is dynamic and changes in response to stimuli, PTMs are important in regulating cellular activity

Common types of Post-translational Modification (PTM)

1. Lipidation
2. Phosphorylation
3. Glycosylation
4. Acetylation
5. Ubiquitination 泛素化
6. Methylation
7. Others

Phosphorylation

Kinases add phosphate groups for activation or inactivation.

Phosphatases hydrolyze the phosphate group (i.e. remove it).

Phosphorylation, principally on serine, threonine or tyrosine* residues, is is critical in the regulation of many cellular processes including cell cycle, growth, apoptosis and signal transduction pathways

* REM: Ser. Thr and Tyr all have –OH in the side chain

Proteins 1

Learning outcome

1. Know basic functions performed by proteins
2. Learn about composition of proteins
3. Recognize the structure of amino acids & understand their basic properties
4. Explain the structure of peptide bond

Proteins 2

Learning objectives

• Understand the forces that hold tertiary structures together
• Understand how globular and membrane bound proteins differ in their structure
• Using myoglobin and haemoglobin as examples understand the significance of protein structure and function
• Know the general features
• Understand how enzymes speed up a reaction
• Understand enzyme diseases

FORCES HOLDING STRUCTURES

1. Covalent
2. Weak forces
   1. Hydrogen bonds
   2. van der Waals forces
   3. Ionic and electrostatic forces

Covalent

Strongest side chain interaction

Hydrogen bonds

Most important inside protein where protected from exchange with solvent water molecules

van der Waals forces

Electronic charge in atom is not evenly distributed all the time.

Atoms attract one another (transient induced dipoles) until they reach Van der Waals distance and then they repel.

When large molecules approach each other many atoms come into contact and net effect is the sum of many atom pairs

Ionic and electrostatic forces

Two oppositely charged molecules attract. Can be stronger in non-polar parts of protein

MYOGLOBIN (Mb)

Found in skeletal muscle MYOGLOBIN

Is the O2 storage protein

78% a-helices (8 segments A to H) linked by turns & no b-sheets

Folded tertiary structure: hydrophobic residues compacted inside, hydrophilic R groups mainly outside. Prolines mainly in bends

Haemoglobin (Hb)

Found in blood packaged inside erythrocytes
Carries O2
Has 4 subunits – a tetramer (quaternary structure)
Each subunit has a haem group
Two types of subunit, called α and ß
α and ß differ in sequence

Mb and Hb have similar 3D folds

The 3D fold of myoglobin can overlay that of a subunit of haemoglobin even though their sequences are very different

You can get the same fold in protein structure from different sequences.

Physical transmission of information

When the O2 binds to the Fe it forces it back into the plane, also pulling the helix

The pull on helix F then moves the EF & the FG corners

This movement reaches αß interface & is transmitted by helix movement over the next haem

It then eases movement of next Fe into haem plane assisting second O2 to bind

On binding the first O2 the dimer α1ß1 and α2ß2 rotates with respect to each other by 15° This causes a change in quaternary structure.

Enzymes

General Features of enzymes

1. They are usually proteins.
2. Enzymes tend to be fairly large molecules usually 104 – 106 daltons
3. Only a relatively small part of whole 3D structure is actually used to catalyse (the active site)
4. Some enzymes have allosteric sites (remote from active site); ligands binding to these sites control rate and/or specificity

EBL Case: Colorectal Cancer

1. There are known factors that increase your risk of developing colorectal cancer.
   Some of these cannot be changed and can be associated with genes, ethnicity (non-modifiable) and demographics such as age. Others can relate to behaviours and our lifestyles – actions we chose to take - and can be modified.
   What risk factors associate with cancer?
   Are there behaviour/lifestyle changes that could be advised to reduce risk?

2. Cancer cells behave differently to normal cells. The difference have been termed “the hallmarks of cancer”.
   List three cancer hallmarks.
   For each one describe how it gives rise to the associated cancer characteristic

3. There are specific genes that are frequently mutated in colorectal cancer.
   These include mutations in tumour suppressor genes encoding APC and p53 and oncogenes encoding KRAS and BRAF.
   Describe the key differences between tumour suppressor and oncogenes. How are they changed in cancer?
   Select one of the listed tumour suppressor proteins. Describe how it works, what happens in colorectal cancer and how this affects the cancer hallmarks.
   Select one of the listed oncoproteins. Describe how it works, what happens in colorectal cancer and how this affects the cancer hallmarks.

4. Describe the cell cycle and how a normal cell becomes a cancer cell.
   What are the key points of regulation?
   How are these changed in cancer?
   What kinds of proteins are involved?

5. The patient has been prescribed chemotherapy - oxaliplatin and capecitabine -in the trigger case above.
   What is the mechanism of action of these agents?
   How do they cause anti-cancer effects?

6. The patient has had a pharmacogenomic screen to see whether they have a variant form of the DPYD gene that would encode a DPYD protein with reduced function.
   Why is it important to know the patient’s DPYD status before administering capecitabine?
   How would DPYD status influence Capacitabine dosing?
   (Referring to your pharmacokinetic lectures may help with these questions)

   

   

7. What are the key symptoms that a person may discuss with a pharmacist that would raise the pharmacists concern that the person may need to be referred for colorectal cancer tests?
   (In the UK, symptoms that would raise concern for a particular condition are known as the “red flag symptoms”).