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Molecular Biology and Public Health （分子生物学与公共卫生）

-----------basic molecules of life（生命的基本分子）

The origin of life molecules

• CH4, H2,NH3,H2O, CO2

• Lightning

• HCN (氰氢酸)• Urea (尿素)• Adenine (腺嘌呤)

Nucleobases（核酸碱基） and Nucleosides（核苷）

• 5 bases: The primary nucleobases are cytosine(胞嘧啶，DNA and RNA), guanine(鸟嘌呤，DNA and RNA), adenine(腺嘌呤，DNA and RNA), thymine (胸腺嘧啶，DNA) and uracil (尿嘧啶，RNA), abbreviated as C, G, A, T, and U, respectively.

• Ribose （核糖）• Phosphate groups （磷酸基团）• Nucleoside（核苷酸）, oligonucleotides （寡核苷酸）

Nucleotide（核苷酸）

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Amino acids（氨基酸）• Amino acids are biologically important organic compounds made

from amine (-NH2) and carboxylic acid (-COOH) functional groups, along with a side-chain specific to each amino acid.

• The key elements of an amino acid are carbon, hydrogen, oxygen, and nitrogen, though other elements are found in the side-chains of certain amino acids.

• 甘氨酸（Glycine）、丙氨酸（Alanine）、缬氨酸（Valine）、亮氨酸（Leucine）、异亮氨酸（Isoleucine）、苯丙氨酸（Phenylalanine）、色氨酸（Tryptophan）、酪氨酸（Tyrosine）、天冬氨酸（Aspartic acid）、组氨酸（Histidine）、天冬酰胺（Asparagine）、谷氨酸（Glutamic acid）、赖氨酸（Lysine）、谷氨酰胺（Glutamine）、甲硫氨酸（Methionine）、精氨酸（Arginine）、丝氨酸（Serine）、苏氨酸（Threonine）、半胱氨酸（Cysteine）、脯氨酸（Proline）等

• 9 of the 20 standard amino acids are called "essential" for humans because they cannot be created from other compounds by the human body, and so must be taken in as food.

Peptides（肽） and Proteins（蛋白质）

• Peptides are short chains of amino acid monomers linked by peptide (amide) bonds.

• Proteins consist of one or more polypeptides arranged in a biologically functional way, often bound to ligands such as coenzymes and cofactors, or to another protein or other macromolecule (DNA, RNA, etc.), or to complex macromolecular assemblies.

• Peptides are distinguished from proteins on the basis of size, and as a benchmark can be understood to contain approximately 50 amino acids or less.

Two basic characters of life: metabolism（代谢） and

reproduction （生殖/繁殖）

• Metabolism is the set of life-sustaining chemical transformations within the cells of living organisms.

• Reproduction (or procreation) is the biological process by which new "offspring" individual organisms are produced from their "parents". Reproduction is a fundamental feature of all known life; each individual organism exists as the result of reproduction. The known methods of reproduction are broadly grouped into two main types: sexual and asexual.

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RNA（核糖核酸）

• Ribonucleic acid (RNA) is a ubiquitous family of large biological molecules that perform multiple vital roles in the coding, decoding, regulation, and expression of genes.

• mRNA, rRNA, tRNA, tmRNA, microRNA, antisense RNA, Ribozyme

mRNA• Messenger RNA (mRNA) is a large family of

RNA molecules that convey genetic information from DNA to the ribosome, where they specify the amino acid sequence of the protein products of gene expression.

• Following transcription of mRNA by RNA polymerase, the mRNA is translated into a polymer of amino acids: a protein, as summarized in the central dogma of molecular biology.

The "life cycle" of an mRNA in a eukaryotic cell. RNA is transcribed in the nucleus; processed, it is transported to the cytoplasm and translated by the ribosome. Finally, the mRNA is degraded.

rRNA• Ribosomal ribonucleic acid (rRNA) is the RNA component of the

ribosome, and is essential for protein synthesis in all living organisms.

• Ribosomes contain two major rRNAs and 50 or more proteins. The LSU and SSU rRNAs are found within the large and small ribosomal subunits, respectively. The LSU rRNA acts as a ribozyme, helping catalyze peptide bond formation.

40S (18S: 1869 nt)60S (5S: 121 nt, 5.8S: 156 nt, 28S: 5070 nt)80Seukaryotic

30S (16S: 1542 nt)50S (5S: 120 nt, 23S: 2906 nt)70Sprokaryotic

Small subunit(rRNA)Large subunit (rRNAs)SizeType

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tRNA• A Transfer RNA (tRNA) is an adaptor molecule

composed of RNA, typically 73 to 94 nucleotides in length, that serves as the physical link between the nucleotide sequence of nucleic acids (DNA and RNA) and the amino acid sequence of proteins.

• It does this by carrying an amino acid to the protein synthetic machinery of a cell (ribosome) as directed by a three-nucleotide sequence (codon) in a messenger RNA (mRNA). As such, tRNAs are a necessary component of protein translation, the biological synthesis of new proteins according to the genetic code.

The interaction of tRNA and mRNA in protein synthesis.

tmRNA

• Transfer-messenger RNA (abbreviated tmRNA, also known as 10Sa RNA and by its genetic name SsrA) is a bacterial RNA molecule with dual tRNA-like and messenger RNA-like properties. The tmRNA forms a ribonucleoprotein complex (tmRNP) together with Small Protein B (SmpB), Elongation Factor Tu (EF-Tu), and ribosomal protein S1. tmRNA combines features of tRNA and mRNA.

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trans-Translation stages A through F. A ribosome with its RNA binding sites, designated E, P, and A, is stuck near the 3' end of a broken mRNA. The tmRNP binds to the A-site, allowing the ribosome to switch templates from the broken message onto the open reading frame of the tmRNA via the resume codon (blue GCA). Regular translation eventually resumes. Upon reaching the tmRNA stop codon (red UAA), a hybrid protein with a proteolysis tag (green beads) is released.

microRNA• A microRNA (abbr. miRNA) is a small non-coding RNA

molecule (ca. 22 nucleotides) found in plants and animals, which functions in transcriptional and post-transcriptional regulation of gene expression.

• Encoded by eukaryotic nuclear DNA, miRNAs function via base-pairing with complementary sequences within mRNA molecules, usually resulting in gene silencing via translational repression or target degradation.

• The human genome may encode over 1000 miRNAs, which may target about 60% of mammalian genes and are abundant in many human cell types.

miRNA Processing Pathway. (1) miRNAs are expressed in the nucleus as parts of long primary miRNA transcripts (Pri-miRNA) that have 5’ caps and 3’poly(A) tails. (2) The hairpin structure that likely forms around the miRNAsequence of the pri-miRNA acts as a signal for digestion by a double-stranded (ds) ribonuclease (Drosha) to produce the precursor miRNA (Pre-miRNA). (3)Exportin-5 mediates nuclear export of pre-miRNAs. (4) A cytoplasmic dsRNAnuclease (Dicer) cleaves the pre-miRNA leaving 1–4 nt 3' overhangs. The single-stranded mature miRNA associates with a complex that is similar, if not identical, to the RNA Induced Silencing Complex (RISC). (5) The miRNA/RISC complex represses protein translation by binding to sequences in the 3' untranslated region of specific mRNAs.

antisense RNA• Antisense RNA (asRNA) is a single-stranded

RNA that is complementary to a messenger RNA (mRNA) strand transcribed within a cell. Some authors have used the term micRNA(mRNA-interfering complementary RNA) to refer to these RNAs but it is not widely used.

• Antisense RNA may be introduced into a cell to inhibit translation of a complementary mRNA by base pairing to it and physically obstructing the translation machinery. This effect is therefore stoichiometric.

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Ribozyme• A ribozyme (ribonucleic acid enzyme) is an RNA

molecule that is capable of performing specific biochemical reactions, similar to the action of protein enzymes.

• The 1981 discovery of ribozymes demonstrated that RNA can be both genetic material (like DNA) and a biological catalyst (like protein enzymes), and contributed to the RNA world hypothesis, which suggests that RNA may have been important in the evolution of prebiotic self-replicating systems.

The chicken-egg debate: RNA world （RNA世界假说）

• The RNA world hypothesis proposes that self-replicating ribonucleic acid (RNA) molecules were precursors to current life, which is based on deoxyribonucleic acid (DNA), RNA and proteins.

• RNA stores genetic information like DNA, and catalyzes chemical reactions like an enzyme protein. It may, therefore, have played a major step in the evolution of cellular life.

DNA（脱氧核糖核酸）

• Deoxyribonucleic acid (DNA) is a molecule that encodes the genetic instructions used in the development and functioning of all known living organisms and many viruses.

• Along with RNA and proteins, DNA is one of the three major macromolecules essential for all known forms of life.

• The structure of the DNA double helix（DNA双螺旋结构）

• Plasmid DNA, chromosome DNA, cDNA, gene (ORF) structure, DNA methylation, chromosome structure

DNA Structure

• Base pairing

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Double helix• The DNA double helix is

stabilized primarily by two forces: hydrogen bonds between nucleotides and base-stacking interactions among aromatic nucleobases.

• The double-helix model of DNA structure was first published in the journal 《Nature》 by James D. Watson and Francis Crick in 1953

DNA双螺旋结构模型的建立

诺贝尔医学与生理学奖1962年

Chromosome （染色体）

• A chromosome is a single piece of coiled DNA containing many genes, regulatory elements and other nucleotide sequences.

• Also contain DNA-bound proteins, which serve to package the DNA and control its functions.

Plasmid（质粒） DNA• A plasmid is a small DNA molecule that is physically

separate from, and can replicate independently of, chromosomal DNA within a cell.

• Most commonly found as small circular, double-stranded DNA molecules in bacteria, plasmids are sometimes present in archaea and eukaryotic organisms.

• In nature, plasmids carry genes that may benefit survival of the organism (e.g. antibiotic resistance), and can frequently be transmitted from one bacterium to another (even of another species) via horizontal gene transfer.

• Artificial plasmids are widely used as vectors in molecular cloning, serving to drive the replication of recombinant DNA sequences within host organisms.

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Plasmid（质粒） DNA Complementary DNA• Complementary DNA (cDNA) is DNA synthesized from a

messenger RNA (mRNA) template in a reaction catalysed by the enzymes reverse transcriptase and DNA polymerase.

• The major difference between cDNA and its corresponding gene sequence is that cDNA has no intron sequences, non-coding DNA sequences punctuated between coding sequences.

Gene（基因）• A modern working definition of a gene is "a

locatable region of genomic sequence, corresponding to a unit of inheritance, which is associated with regulatory regions, transcribed regions, and or other functional sequence regions ".

• ORF: Open reading frame(开放阅读框)• Genetic code(密码子)

DNA codon（密码子）/ Genetic code（遗传编码）

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Chromosome structure• Bacterial Chromosome Structure

Prokaryotic cells (bacteria) contain their chromosome as circular DNA. Usually the entire genome is a single circle, but often there are extra circles called plasmids. The DNA is packaged by DNA-binding proteins.

Chromosome structure• Eukaryotic chromosome structure

Eukaryotic cells contain their DNA within the nuclear membrane. The DNA is wrapped around the histone core of eight protein subunits, forming the nucleosome. The nucleosome is clamped by histone H1. About 200 base pairs (bp) of DNA coil around one histone.

DNA methylation (甲基化)• DNA methylation is a biochemical process involving the

addition of a methyl group to the cytosine or adenine DNA nucleotides.

• DNA methylation stably alters the expression of genes in cells as cells divide and differentiate from embryonic stem cells intospecific tissues.

• The resulting change is normally permanent and unidirectional, preventing one organism from reverting to a stem cell or converting into another type of tissue.

•Methylation of histone or of DNA usually turns a gene off.•Acetylation of histone usually turns a gene on.•Phosphorylation -- we're not sure what that does.

Gene expression flow

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Peptides • Peptides are short chains of amino acid monomers

linked by peptide (amide) bonds, the covalent chemical bonds formed when the carboxyl group of one amino acid reacts with the amino group of another. Peptides are distinguished from proteins on the basis of size, and as a benchmark can be understood to contain approximately 50 amino acids or less

Single unit proteins（单链蛋白质 ）

• Contains only one peptide

• Bacterial proteins

• Ribonuclease A

• lysozyme

Multi-unit proteins（寡聚蛋白质）

• Contains 2 or >2 peptides （subunits）

• Eukaryotic proteins

Protein complexes 蛋白复合体

• Contains 2 or >2 proteins

• Linked by non-covalent protein–proteininteractions

• These complexes are a cornerstone of many (if not most) biological processes and together they form various types of molecular machinery that perform a vast array of biological functions.

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Central dogma of molecular biology (中心法则)

• The central dogma of molecular biology is an explanation of the flow of genetic information within a biological system.

• It was first stated by Francis Crick in 1958: “The central dogma of molecular biology deals with the detailed residue-by-residue transfer of sequential information. It states that such information cannot be transferred from protein to either protein or nucleic acid”.

• Marshall Nirenberg said, "DNA makes RNA makes protein.“

Genetic information flow

The exceptions (deviations) of central dogma landed two Nobel prizes

David Baltimore in 1975 for his discovery of Reverse transcriptase (RT)

Stanley Benjamin Prusiner in 1997 for his discovery of prion.

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DNA RNA Protein

Summary• Life molecule: DNA, RNA, Protein

• RNA: mRNA, rRNA, tRNA, tmRNA, microRNA, antisense RNA, Ribozyme

• DNA: structure, plasmid DNA, chromosome, cDNA, gene (ORF) structure, DNA methylation, chromosome structure

• Peptides, single unit proteins, multi-unit proteins, protein complexes

• Central dogma of molecular biology

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Questions

1. Why RNA is the most likely original molecule of life?

2. Why cells do not use RNA alone as basic life molecules despite that RNA can both store genetic information and catalyze biochemical reactions?

3. What is the core definition of central dogma and how it has been challenged and supplemented?

The mission of scientist

Uncover the law and truth of mother nature

Persist (truth often lies in the hands of minority)

Challenge authority (Respect law of nature more than respect any person

Comprehensive (no rush to conclusion, self-correct)

Curiosity and unlimited imagination

Suggested reading: “The structure of Scientific Revolutions by Thomas S. Kuhn

A quiz for fun

1. Which month has 28 days?

2. What can’t you ever grab with your right hand?

3. How many eggs can you eat with an empty stomach?

4. Can you imagine something that might travel faster than light?

Next…

• DNA manipulation: DNA Isolation, gel electrophoresis, Restriction enzymes (scissors), ligases, polymerases, polynucleotide kinases, TDT, DNA topoisomerases, vectors, amplification, Southern blot, primer extension, RFLP, PFGE, Multi-locus sequencing, Whole genome sequencing, comparative genomics