Understanding Proteins and Nucleic Acids: From Amino Acids to DNA Replication, Study notes of Chemistry

Download Understanding Proteins and Nucleic Acids: From Amino Acids to DNA Replication and more Study notes Chemistry in PDF only on Docsity! Today’s plan - A few minutes with amino acids and proteins. Then, all of chapter 3, “From Genes to Proteins”. Proteins are polymers of amino acids, linked together by “peptide bonds”. Single amino acid. “R” group is different, depending on a.a. type. 20 different amino acid types commonly occur in proteins. Examples of proteins: Myoglobin is involved in oxygen transport. Keratin is a fibrous protein, found in hair, finger nails, tortoise shells, etc. Protein function depends on its structure; Structure depends on the amino acid sequence. Amino acids. Either way is OK. Textbook likes this.I like to draw them like this. Structures and abbreviations of the 20 standard amino acids Hydrophobic amino acids (or non-polar) coo coo coo i Wey noe wc-an-X Y \—; NH} NHS NH} NHS Alanine (Ala, A) Valine (Val, V) Phenylalanine (Phe, F) Tryptophan (Trp, W) T T ee Ta | a) 1—( SS — NH NHS NHS HNL Leucine (Leu, L) Isoleucine (Ile, |) Methionine (Met, M) Proline (Pro, P) Polar amino acids (or hydrophilic) coo f° f° r : nt emeson Hd n-c-on{ Yon wt NHS NHS NHS NH} Serine (Ser, S) Threonine (Thr, T) Tyrosine (Tyr, ¥) Cysteine (Cys, C) re Te rT am ns "—< HoH NH; NH NH; NH Asparagine (Asn, N) Glutamine (Gin, Q) Histidine (His, H) Glycine (Gly, G) Charged amino acids Te j T ° f T. NH, me h—C—O- 1 —o- 1 ee HEH, CH, NC NHy NH} NH; NH; Aspartate (Asp, D) Glutamate (Glu, E) Lysine (Lys, K) Arginine (Arg, R) A few words about DNA & RNA. These are polymers of nucleic acids. Components of DNA & RNA: DNA: adenine (A), guanine (G), cytosine (C), thymine (T). RNA: adenine (A), guanine (G), cytosine (C), uracil (U). A & G have a similar “purine” ring structure. C, T & U have a similar “pyrimidine” ring structure. Uracil (U) is a component of RNA. Looks like thymine, but with methyl group. “Nucleosides” - A base attached to a ribose sugar ring. Ribose (RNA) 2-deoxyribose (in DNA) Some nucleotides of DNA. dAMP (deoxyadenosine monophosphate) dGDP (deoxyguanosine diphosphate) dCTP (deoxycytosine triphosphate) Vocabulary. “NTPs” means nucleotides of RNA all together. NTPs = (ATP + CTP + GTP + UTP) “dNTPs” means nucleotides of DNA all together. dNTPs = (dATP + dCTP + dGTP + dTTP) Structure of a single strand of DNA polymer. sugar-phosphate backbone sugar-phosphate backbones Major and minor grooves in DNA. minor major major groove minor groove Rotate 30 degrees around this axis. RNA can also form base pairs. A = U C = G 2 strands of base paired RNA: RNA can form complex structures: A stem-loop in a viral RNA. A “pseudoknot” within a messenger RNA. 5! fi Central dogma of molecular biology: transcription translation ° Sranseripon, mRNA a" Protein (messenger RNA) DNA “Transcription” is carried out by RNA polymerase. RNA polymerase makes a strand of RNA that is complementary to the bottom strand of a DNA template. RNA polymerase “reads” DNA as “ACGT”, writes RNA as “ACGU”. “Translation” is carried out by a “ribosome” & transfer RNA (tRNA): The ribosome & tRNAs read the mRNA, and make protein using amino acids. & ribosome (RNA polymerase) (ribosome & tRNA)translation transcription DNA sequence codes for mRNA. mRNA codes for protein sequence: Absorbance of UV light by nucleic acids increases by about 30% when the base pairs are melted: From a melting curve such as above, it is possible to determine the “melting temperature” of nucleic acids. 1) Detecting “melting” of DNA by change in absorbance of UV light. Absorbance of UV light at 260 nm Temperature (deg C) Why is there a change in UV absorbance upon melting DNA (or RNA)? A change in absorbance (at 260 nm) that is associated with changes in base stacking. Remember Beer’s law: A =€C “e” for nucleic acids is lower when base stacking occurs (because base stacking changes the energies of the 1 -> m* electronic transitions that give rise to the absorbance at 260 nm). DNA sequencing uses DNA polymerase and dNTPs with a little “dideoxy-NTPs” (ddNTPs) mixed in. When the DNA pol uses a ddNTP, the polymerization stops. The resulting DNA ends with a “ddNTP” nucleotide, which can not be extended further. 2) DNA sequencing. Figure from the text, showing DNA strands of various lengths migrating through an electric field. The color identifies the type (A, C, G or T) of the ddNTP that terminated the strand. The whole human genome was sequenced a few years ago! E. coli bacteria have about 4000 genes for 4000 different proteins. What things do you need to do PCR? DNA template (to be copied) & DNA short primers. dNTPs (mixed dATP, dCTP, dGTP, dTTP) DNA polymerase (preferable thermostable, so it can stand the heating and cooling cycles) A popular DNA polymerase for use in PCR: DNA pol from Thermus aquaticus (“Taq polyerase”) DNA pol from Pyrococcus furiosus (“Pfu polymerase”, from a thermophilic archeaon found near Vulcano Island, Italy) (figure at end of chapter 3) The polymerase chain reaction. —_ oO 1 “ { a \ \ urate sranis  More tools of nucleic acid chemistry: 3) Restriction enzymes: Cut DNA at specific sequences. Maybe move this to proteins lecture 2 - add section on “recombinant protein” Production of a recombinant DNA molecule. cut ~@" () O ligate O —_— Recombinant DNA Plasmid Foreign DNA  Hypochromism in nucleic acids. Defined as a change in absorbance (at 260 nm) that is associated with changes in base stacking. Remember Beer’s law: A = ε C “ε” for nucleic acids is lower when base stacking occurs (because base stacking changes the energies of the π -> π* electronic transitions that give rise to the absorbance at 260 nm). Hypochromism in nucleic acids. Defined as a change in absorbance (at 260 nm) that is associated with changes in base stacking. Remember Beer’s law: A = ε C “ε” for nucleic acids is lower when base stacking occurs (because base stacking changes the energies of the π -> π* electronic transitions that give rise to the absorbance at 260 nm). Hypochromism provides a means for detecting structural changes in nucleic acids. For example: From a “melting curve” such as above, it is possible to quantitatively evaluate nucleic acid stability. Normal gene 504 505 506 507 508 509 510 511 512 mRNA *** GAA AAT ATC ATC TTT GGT GTT TCC TAT ::: Protein **+ Glu Asn Ile tle Phe Gly Val Ser Tyr -:- Mutated gene 504 505 506 507 508 509 510 511 512 mRNA *** GAA AAT ATC AT- --T GGT GTT TCC TAT ::: Protein **+ Glu Asn Ile Ile Gly Val Ser Tyr -:: One of the abnormalities found in Cystic Fibrosis Unnumbered, page 69 (2) Deletion.  Transcription as described in the textbook: Coding strand (nontemplate) Noncoding strand (template)