Protein Biochemistry
Protein Functions |
Structure |
Post-translational modification & Targeting |
Different structures reflect unique function |
Proteins are made up of amino acids with various side chains |
Reversible (addition) or irreversible (removal) |
Recognition of specific molecules: hormones, antibodies, DNA binding proteins |
Amino acids have a hydrogen, central carbon, amino group, side chain, and a carboxyl group |
Methylation is adding a CH3 group (eg histones to regulate gene expression) |
Movement of molecules: porin, ferritin |
Side chains: positive or negative charge, polar or nonpolar, different shapes and sizes |
Glycosylation is adding sugar molecules (eg cell surface proteins) |
Structural functions: components of the cytoskeleton such as microtubules |
Primary structure: order of amino acids in a polypeptide chain, joined by peptide bonds (which are rigid), have a C and N-terminus |
Ubiquitination is adding a 76 amino acid polypeptide which denotes protein is ready to be degraded |
Enzymes: speed up chemical reactions by lowering the activation energy required |
Secondary structure: alpha helix or beta pleated sheet, stabilised by hydrogen bonds |
Phosphorylation is adding PO3 group, regulates enzyme function |
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Tertiary structure: tightly packed 3D structure, noncovalent interactions between side chains |
Targeting is when proteins are transported to where they need to go in a cell |
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Quaternary structure: complex with 2 or more subunits which can be identical or different |
Many proteins have a short signal or localisation sequence indicating where they need to go, this is then removed |
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Many proteins contain several different tightly packed domains, each carries out a specific function |
DNA Structure
DNA Structure |
Experimental Evidence |
Chromosome Structure |
DNA- Binding Proteins |
DNA is made up of nucleotides |
Chargaff used paper chromatography and looked at base proportions. % purine = % pyrimidine |
Chromosomes are long DNA molecules containing genetic information, have regulatory sequences for proper expression and replication |
Proteins bind to specific domains which can have a general affinity for DNA, or are sequence specific |
Nucleotides have: deoxyribose ring, nitrogenous base, phosphate group |
Wilkins and Franklin used X-Ray crystallography, found DNA is a helix with even structure |
Eukaryotic chromosomes are linear, have a; centromere, and telomeres |
Transcriptional regulators bind regulatory sequences near promoters to block or stimulate transcription (eg lac operon in E.coli) |
Purines (adenine, guanine) have 2 rings, pyrimidines (cytosine, thymine, uracil) have 1 ring |
Watson and Crick made a model: A-T and G-C hydrogen bonded base pairs, antiparallel strands, right handed double helix, one helical turn every 10.5 base pairs (3.4 nm), major and minor grooves |
Bacteria have a smaller single circular chromosome |
Restriction endonucleases are enzymes that cut DNA at specific sequences. Bacteria use them to restrict virus action, they can be used in the lab to manipulate DNA |
DNA is written from 5’ to 3’ |
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Plasmids in prokaryotes can be passed between cells via conjugation |
Histones are proteins that DNA wraps around to form chromatin. Not sequence specific |
2 H bonds between adenine and thymine, 3 H bonds between cytosine and guanine |
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DNA as Genetic Material
Chromosomal Inheritance |
Transforming Principle |
Hershey-Chase Experiment |
Sutton & Boveri investigated where genetic material is carried using cytology, and microscopy |
Griffith worked on S. pneumoniae; S strain are pathogenic (have capsule), R strain is not |
Bacteriophage T2 inject genetic material inside E.coli, investigated what this material is |
Sutton used grasshoppers, Boveri used Ascaris worms (roundworms). Their chromosomes are large and few in number, making them easy to observe |
When cell extract of dead S strain is injected to mice- no illness. When combined with live R strain and injected- illness |
Labelled bacteriophage with radioactive isotopes. 32P for DNA, 35S for protein to deduce which is genetic material |
Discovered chromosomes are important in reproduction and development |
Bacteria are being transformed when combined, hereditary material is being passed |
Allowed bacteriophage to inject unlabelled bacteria. Separated phage from bacteria using blender |
Discoveries matched those of Mendel's, and provided physical basis for his theories |
Tested which molecule carries hereditary material, used enzymes which destroy specific molecules. |
Centrifuged. Tested infected bacteria pellet with Geiger counter. |
Suggested different combinations of chromosomes could cause variation; discovered genes, and the linear structure of chromosomes |
Discovered DNA is responsible for transformation. Gene coding for the capsule is passed to R strain from S strain, making them pathogenic |
Bacteriophage labelled 32P had made the bacteria radioactive, indicating DNA is genetic material |
DNA Replication
Semi- Conservative Replication |
Process of Replication |
Enzymes for Replication |
Leading and Lagging Strands & Telomeres |
DNA strands are complementary |
DNA strands separate and are used as templates for new strands |
Polymerase adds nucleotides in a 5’ to 3’ direction, needs primer to start |
Leading strand is 5’ to 3’, while the lagging strand is 3’ to 5’ direction |
3 theories for replication: conservative, semi-conservative, dispersive |
Replication fork- region where DNA is being copied |
Primase generates primer (usually RNA), a small stretch of nucleotides in a 5’ to 3’ direction. Removed afterwards and the gap is filled in (by polymerase) |
Replication in lagging strand leads away from fork and is discontinuous. Strand is primed many times, so Okazaki fragments form. |
Meselson- Stahl used nitrogen isotopes to test which theory is correct. Grew E.coli in 15N (to make heavy DNA) and transferred to 14N |
Origin of replication- where the hydrogen bonds are broken and the strands are pulled apart so replication can start |
Single stranded binding proteins separate the DNA strands and prevent reannealing |
Primer removal at the end of Okazaki fragments causes erosion of genetic material, telomeres solve this |
Separated heavy and light DNA by ultracentrifugation, obtained a liquid gradient. |
Humans have multiple origins of replication, E.coli have one |
Helicase breaks the hydrogen bonds between bases and unwinds the helix |
Telomeres- short stretches of repetitive DNA sequences at the end of chromosomes, some is lost after replication |
Observed using UV light, after 1 generation DNA was hybrid. After 2+ generations it became lighter, proving semi-conservative replication |
Replication is bidirectional |
Ligase joins the stretches of DNA together into a single strand |
Telomeres are effective where DNA needs to be passed on perfectly |
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Topoisomerase relieves pressure from overwinding around the replication bubble by making and resealing breaks in the DNA |
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