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04/19/23 dmitra 1
Molecular Biology Background
Debasis MitraFlorida Tech
Credit: Pevezner text-site
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Section1: What is Life made of?
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2 types of cells: Prokaryotes v.s.Eukaryotes
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Life begins with Cell
A cell is a smallest structural unit of an organism that is capable of independent functioning
All cells have some common features
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Prokaryotes and Eukaryotes
•According to the most recent evidence, there are three main branches to the tree of life. •Prokaryotes include Archaea (“ancient ones”) and bacteria.•Eukaryotes are kingdom Eukarya and includes plants, animals, fungi and certain algae.
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Prokaryotes and Eukaryotes, continued
Prokaryotes Eukaryotes
Single cell Single or multi cell
No nucleus Nucleus
No organelles Organelles
One piece of circular DNA
Chromosomes
No mRNA post transcriptional modification
Exons/Introns splicing
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Overview of organizations of life Nucleus = library Chromosomes = bookshelves Genes = books Almost every cell in an organism
contains the same libraries and the same sets of books.
Books represent all the information (DNA) that every cell in the body needs so it can grow and carry out its vaious functions.
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Chromosomes
Organism Number of base pair number of Chromosomes
---------------------------------------------------------------------------------------------------------
Prokayotic
Escherichia coli (bacterium) 4x106 1
Eukaryotic
Saccharomyces cerevisiae (yeast) 1.35x107 17
Drosophila melanogaster(insect) 1.65x108 4
Homo sapiens(human) 2.9x109 23
Zea mays(corn) 5.0x109 10
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Bio-molecules
Nucleic acids (DNA, RNA): Library of life
Proteins: Workhorse of life Fatty acids, carbohydrates, and other
supporting molecules
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DNA DNA has a double helix
structure which composed of sugar molecule phosphate group and a base (A,C,G,T)
DNA always reads from 5’ end to 3’ end for transcription replication 5’ ATTTAGGCC 3’3’ TAAATCCGG 5’
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DNA, RNA, and the Flow of Information
TranslationTranscription
Replication
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Proteins Functions
Structural Enzymes Information exchange (e.g., across
cell walls) Transporting other molecules (e.g.,
oxygen to cells) Activating-deactivating genes Etc.
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Proteins
Amino acids
Protein is a chain of “residues”
20 to 5000 long, typically a few hundred long
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Protein structure
Important for its function Primary structure: sequence Secondary structure: a few
topological features Tertiary structure: 3D folding Quaternary structure: Protein
complex
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Protein Folding Proteins tend to fold into the lowest free
energy conformation. Proteins begin to fold while the peptide is
still being translated. Proteins bury most of its hydrophobic
residues in an interior core to form an α helix.
Most proteins take the form of secondary structures α helices and β sheets.
Molecular chaperones, hsp60 and hsp 70, work with other proteins to help fold newly synthesized proteins.
Much of the protein modifications and folding occurs in the endoplasmic reticulum and mitochondria.
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Protein Folding (cont’d)
The structure that a protein adopts is vital to it’s chemistry
Its structure determines which of its amino acids are exposed carry out the protein’s function
Its structure also determines what substrates it can react with
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Nucleic acids
Two types:
DNA: Deoxy-ribonucleic acid RNA: Ribonucleic acid
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Nucleic acids Sugar molecule chain forms the
base of the polymer
Two types of sugar: ribose (RNA), 2’-deoxyribose (DNA)
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Nucleic acids: DNA
4 types of bases connected to sugar molecules: Adenine (a), Guanine (g), Thymine (t) and Cytosine (c)
A and T forms strong bonds, and so do G and C
An Introduction to Bioinformatics Algorithms www.bioalgorithms.info
04/19/23 2015
The Purines The Pyrimidines
An Introduction to Bioinformatics Algorithms www.bioalgorithms.info
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DNA• DNA has a double helix
structure which composed of • sugar molecule• phosphate group• and a base (A,C,G,T)
• DNA always reads from 5’ end to 3’ end for transcription replication 5’ ATTTAGGCC 3’3’ TAAATCCGG 5’
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Nucleic acids: DNA
Double stranded: two strands of sugar molecule-chains
Each strand is directed: 5’ to 3’
Attached inside by base-pairings (a-t and g-c)
An Introduction to Bioinformatics Algorithms www.bioalgorithms.info
04/19/23 2315
Double helix of DNA
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Discovery of DNA
DNA Sequences Chargaff and Vischer, 1949
DNA consisting of A, T, G, C• Adenine, Guanine, Cytosine, Thymine
Chargaff Rule Noticing #A#T and #G#C
• A “strange but possibly meaningless” phenomenon.
Wow!! A Double Helix Watson and Crick, Nature, April 25, 1953
Rich, 1973 Structural biologist at MIT. DNA’s structure in atomic resolution.
Crick Watson1 Biologist1 Physics Ph.D. Student900 wordsNobel Prize
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Nucleic acids: DNA
Each strand is complementary and reverse to the other
If s=agacgt
reverse(s)=tgcaga
reverse-complement(s)=acgtct
Double-strand: 5’--agacgt->3’
3’<-t ctgca—5’
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Nucleic acids: DNA 3D structure is helical
Double-stranded helix: like step ladder
Each unit is a base pair (sugar-base-base-sugar)
DNA’s in cells are chromosomes (human chromosome ~3*(10^9) bp long)
Squeezed 3D structure in cell may have functional importance – not well studied
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DNA Replication
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Nucleic acids: RNA
Replace t with u (uracil) as base
May or may not be (mostly not) double stranded
Functions: Information storage like DNA, sometimes workhorse like proteins
Possible evolutionary precursor to DNA and protein
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Genetic code
Proteins do almost all the works!!
Information for coding proteins are stored on DNA’s (or RNA’s): genes
Three consecutive bases on a gene codes an amino acid, or the STOP code: codon
The table is called genetic code
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Cell Information: Instruction book of Life
DNA, RNA, and Proteins are examples of strings written in either the four-letter nucleotide of DNA and RNA (A C G T/U)
or the twenty-letter amino acid of proteins. Each amino acid is coded by 3 nucleotides called codon. (Leu, Arg, Met, etc.)
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Overview of DNA to RNA to Protein
A gene is expressed in two steps
1) Transcription: RNA synthesis2) Translation: Protein synthesis
An Introduction to Bioinformatics Algorithms www.bioalgorithms.info
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Central Dogma of Biology
The information for making proteins is stored in DNA. There is a process (transcription and translation) by which DNA is converted to protein. By understanding this process and how it is regulated we can make predictions and models of cells.
Sequence analysis
Gene Finding
Protein Sequence Analysis
Assembly
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Transcription
Genes are transcribed to proteins: typically one gene to one protein
Genes are subsequenes on chromosomes started by a promoter region, ended around a stop codon
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Transcription
Steps: DNA is split over gene after
promoter is recognized (may have other regulatory regions upstream)
mRNA is copied from the gene Exons are spliced out from the
mRNA keeping the introns only Ribosome (rRNA and protein
complex) works on mRNA
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Transcription The process of
making RNA from DNA
Catalyzed by “transcriptase” enzyme
Needs a promoter region to begin transcription.
~50 base pairs/second in bacteria, but multiple transcriptions can occur simultaneously
http://ghs.gresham.k12.or.us/science/ps/sci/ibbio/chem/nucleic/chpt15/transcription.gif
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Definition of a Gene
Regulatory regions: up to 50 kb upstream of +1 site
Exons: protein coding and untranslated regions (UTR)1 to 178 exons per gene (mean 8.8)8 bp to 17 kb per exon (mean 145 bp)
Introns: splice acceptor and donor sites, junk DNAaverage 1 kb – 50 kb per intron
Gene size: Largest – 2.4 Mb (Dystrophin). Mean – 27 kb.
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Translation
tRNA are attached to codons on mRNA
On the other end the tRNA attracts appropriate amino acid
Amino acids are zipped up No tRNA for STOP codon Every step is facilitated by
appropriate enzymeCentral Dogma of biology
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Translation, continued Catalyzed by Ribosome Using two different sites,
the Ribosome continually binds tRNA, joins the amino acids together and moves to the next location along the mRNA
~10 codons/second, but multiple translations can occur simultaneously
http://wong.scripps.edu/PIX/ribosome.jpg
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Revisiting the Central Dogma
In going from DNA to proteins, there is an intermediate step where mRNA is made from DNA, which then makes protein This known as The
Central Dogma Why the intermediate step?
DNA is kept in the nucleus, while protein sythesis happens in the cytoplasm, with the help of ribosomes
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The Central Dogma (cont’d)
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Open Reading Frame
Three reading frames in a strand Complementary strand may have
another three frames
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Types of chromosomes
Procaryotes (bacteria, blue algae): circular
Eucaryotes (has nuclear wall): diploid (human has 23 pairs)
Homologous genes and alleles (e.g., human hemoglobin of type A, B, and O)
Haploid chromosomes in Eucaryote sex cells
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DNA Sequencing
A DNA fragment is split at each position starting from one end
Four tubes: one containing molecules ending with G, one with A, one with T and another one with C
Electrophoresis separates each chunk of different size in each tube [page 22]
Information is recombined to sequence the DNA chunk
Can be done for the size of only ~1K bp long chunk
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DNA Sequencing
Human DNA is ~10^9 bp long
Restriction enzyme cuts at restriction sites (a product of genetic engineering) [page 18]
After sequencing, information from fragments need to be recombined to get the broader picture
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DNA Sequencing
Depends on finding restriction site/enzyme for fragmenting DNA of appropriate size
Privately funded Tiger project (Celera now) used heat and vibration to create fragments
Recombining information is no longer trivial because fragment’s location is no longer known
Needed Fragment assembly algorithm
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DNA Sequencing
Needs multiple copies of DNAs Recombinant DNA by biologically
copying them within host organisms
Polymerase Chain Reaction: heat and tear two strands of DNA, then let each strand attract nucleic acids to form double stranded DNA, repeat