Molecular Biology Griffith and Transformation Fredrick Griffith isolated

Molecular Biology

Griffith and Transformation Fredrick Griffith isolated two different strains of pneumonia bacteria from mice and grew them in his lab. The disease-causing strain of bacteria grew into smooth colonies on culture plates. The harmless strain grew into colonies with rough edges.

The Central Dogma of Biology dog·ma ˈdôɡmə/ noun: dogma; plural noun: dogmas 1. A principle or set of principles laid down by an authority as incontrovertibly true. "the Christian dogma of the Trinity“ Origin: mid 16 th century: via late Latin from Greek dogma ‘opinion, ’ from dokein ‘seem good, think. ’ Translate dogma to Use over time for: dogma DNA RNA Protein

Griffith's Experiment 1: Mice were injected with the disease-causing strain of bacteria. The mice developed pneumonia and died.

Experiment 2: Mice were injected with the harmless strain of bacteria. Harmless bacteria (rough colonies) These mice didn’t get sick. Lives

Experiment 3: Griffith killed the disease-causing bacteria. He then injected the heat-killed bacteria into the mice. Heat-killed diseasecausing bacteria (smooth colonies) The mice survived. Lives

Experiment 4: Griffith mixed heat-killed, diseasecausing bacteria with live, harmless bacteria and injected the mixture into the mice. The mice developed pneumonia and died. Heat-killed diseasecausing bacteria (smooth colonies) Harmless bacteria (rough colonies) Live diseasecausing bacteria (smooth colonies) Dies of pneumonia

The heat-killed bacteria passed their disease-causing ability to the harmless strain. Griffith called this process transformation because the harmless strain had changed into the disease-causing strain. Griffith hypothesized that a factor must contain information that could change harmless bacteria into disease-causing ones.

Avery and DNA Oswald Avery determined which molecule was most important for transformation. Avery made an extract from the heat-killed bacteria that they treated with enzymes.

Enzymes to destroy proteins Then lipids Then carbohydrates Then other molecules including RNA. Transformation still occurred. Then DNA - NO transformation Therefore, he concluded that DNA was the transforming factor.

Avery and other scientists discovered that the nucleic acid DNA stores and transmits the genetic information from one generation of an organism to the next.

The Hershey-Chase Experiment Alfred Hershey and Martha Chase studied viruses—nonliving particles smaller than a cell that can infect living organisms.

Bacteriophages A virus that infects bacteria is known as a bacteriophage. Bacteriophages are composed of a DNA or RNA core and a protein coat. Copyright Pearson Prentice Hall

They grew viruses in cultures containing radioactive isotopes of phosphorus-32 (32 P) and sulfur-35 (35 S). If 35 S was found in the bacteria, it would mean that the viruses’ protein had been injected into the bacteria. Bacteriophage with suffur-35 in protein coat Phage infects bacterium No radioactivity inside bacterium

If 32 P was found in the bacteria, then it was the DNA that had been injected. Bacteriophage with phosphorus-32 in DNA Phage infects bacterium Radioactivity inside bacterium

Nearly all the radioactivity in the bacteria was from phosphorus (32 P). Hershey and Chase concluded that the genetic material of the bacteriophage was DNA, not protein.

Chargaff's Rules Erwin Chargaff discovered that: • The percentages of guanine [G] and cytosine [C] bases are almost equal in any sample of DNA. • The percentages of adenine [A] and thymine [T] bases are almost equal in any sample of DNA. Copyright Pearson Prentice Hall

DNA A nucleotide is a monomer of nucleic acids made up of: • Deoxyribose – 5 -carbon Sugar • Phosphate Group • Nitrogenous Base

There are four kinds of bases in DNA: • adenine • guanine • cytosine • thymine

X-Ray Evidence Rosalind Franklin aimed an X-ray beam at concentrated DNA samples and recorded the scattering pattern of the X-rays on film. This gave her information about the structure of DNA.

The Double Helix James Watson and Francis Crick stole Franklin’s data and built a model that explained how DNA carried information and could be copied. Watson and Crick's model of DNA was a double helix, in which two strands were wound around each other.

DNA Double Helix

Watson and Crick discovered that hydrogen bonds can form only between certain base pairs—adenine and thymine, and guanine and cytosine. This principle is called base pairing. DNA Structure Video 18 Things You Should Know

Quiz 8 -1

Avery and other scientists discovered that a. DNA is found in a protein coat. b. DNA stores and transmits genetic information from one generation to the next. c. transformation does not affect bacteria. d. proteins transmit genetic information from one generation to the next.

The Hershey-Chase experiment was based on the fact that a. DNA has both sulfur and phosphorus in its structure. b. protein has both sulfur and phosphorus in its structure. c. both DNA and protein have no phosphorus or sulfur in their structure. d. DNA has only phosphorus, while protein has only sulfur in its structure.

DNA is a long molecule made of monomers called a. nucleotides. b. purines. c. pyrimidines. d. sugars.

Chargaff's rules state that the number of guanine nucleotides must equal the number of a. cytosine nucleotides. b. adenine nucleotides. c. thymine nucleotides. d. thymine plus adenine nucleotides.

In DNA, the following base pairs occur: a. A with C, and G with T. b. A with T, and C with G. c. A with G, and C with T. d. A with T, and C with T.

DNA and Chromosomes In prokaryotic cells, DNA is located in the cytoplasm. Most prokaryotes have a single, circular DNA molecule containing nearly all of the cell’s genetic information. Chromosome E. Coli Bacterium Bases on the Chromosomes

Many eukaryotes have 1000 times the amount of DNA as prokaryotes. Eukaryotic DNA is located in the cell nucleus inside chromosomes. The number of chromosomes varies widely from one species to the next.

Chromosome Structure • Eukaryotic chromosomes contain DNA and protein, tightly packed together to form chromatin. • Chromatin consists of DNA tightly coiled around proteins called histones. • DNA and histone molecules form nucleosomes. • Nucleosomes pack together, forming a thick fiber. Chromosome Nucleosome DNA double helix Coils Supercoils Histones

The two strands in the DNA backbone run in "anti-parallel" directions to each other. That is, one of the DNA strands is built in the 5’ 3’ direction, while the complementary strand is built in the 3’ 5’ direction.

DNA Replication Each strand of the DNA double helix has all the information needed to reconstruct the other half by the mechanism of base pairing. In most prokaryotes, DNA replication begins at a single point and continues in two directions. DNA Replication Video

In eukaryotic chromosomes, DNA replication occurs at hundreds of places. Replication proceeds in both directions until each chromosome is completely copied. The sites where separation and replication occur are called replication forks.

Duplicating DNA Before a cell divides, it duplicates its DNA during “S” phase in a process called replication. Replication ensures that each resulting cell will have a complete set of DNA.

During DNA replication, the DNA molecule separates into two strands, then produces two new complementary strands following the rules of base pairing. Each strand of the double helix of DNA serves as a template for the new strand.

New Strand Original strand Nitrogen Bases Growth Replication Fork DNA Polymerase

How Replication Occurs DNA replication is carried out by enzymes that “unzip” a molecule of DNA. Hydrogen bonds between base pairs are broken and the two strands of DNA unwind. Copyright Pearson Prentice Hall

The principal enzyme involved in DNA replication is DNA polymerase joins individual free nucleotides to produce a DNA molecule and then “proofreads” each new DNA strand.

Quiz 8– 2

In prokaryotic cells, DNA is found in the a. cell membrane. b. nucleus. c. ribosome. d. cytoplasm.

The first step in DNA replication is a. producing two new strands. b. producing DNA polymerase. c. separating the strands. d. correctly pairing bases.

A DNA molecule separates, and the sequence GCGAATTCG occurs in one strand. What is the base sequence on the other strand? a. GCGAATTCG b. GATGGCCAG c. TATCCGGAT d. CGCTTAAGC

In addition to carrying out the replication of DNA, the enzyme DNA polymerase also functions to a. “proofread” the new copies to minimize the number of mistakes. b. regulate the time copying occurs in the cell cycle. c. unzip the DNA molecule. d. wrap the new strands onto histone proteins.

The structure that may play a role in regulating how genes are “read” to make a protein is the a. coil. b. histone. c. chromatin. d. nucleosome.

RNA and Protein Synthesis

• Genes are coded DNA instructions that control the production of proteins. • Genetic messages can be decoded by copying part of the nucleotide sequence from DNA into m. RNA. • m. RNA contains coded information for making proteins.

The Structure of RNA There are three main differences between RNA and DNA: • The sugar in RNA is ribose instead of deoxyribose. • RNA is generally single-stranded. • RNA contains uracil in place of thymine.

Types of RNA There are three main types of RNA: • messenger RNA (m. RNA) • ribosomal RNA (r. RNA) • transfer RNA (t. RNA) Transcription & Translation Video

Messenger RNA (m. RNA) carries copies of instructions for assembling amino acids into proteins.

Ribosome Ribosomal RNA Ribosomes are made up of proteins and ribosomal RNA (r. RNA).

Amino acid Transfer RNA During protein construction, transfer RNA (t. RNA) transfers each amino acid to the ribosome.

DNA molecule DNA strand (template) 5¢ 3¢ TRANSCRIPTION m. RNA 5¢ 3¢ Codon TRANSLATION Protein Amino acid

Transcription DNA is copied in the form of RNA This first process is called transcription. The process begins at a section of DNA called a promoter.

RNA Editing Some DNA within a gene is not needed to produce a protein. These areas are called introns. The DNA sequences that code for proteins are called exons.

The introns are cut of RNA molecules. Exon Intron DNA Pre-m. RNA The exons are the spliced together to form m. RNA Cap Tail

The Genetic Code The genetic code is the “language” of m. RNA instructions. The code is written using four “letters” (the bases: A, U, C, and G).

A codon consists of three consecutive nucleotides on m. RNA that specify a particular amino acid.

Translation is the decoding of an m. RNA message into a polypeptide chain (protein). Translation takes place on ribosomes. During translation, the cell uses information from messenger RNA to produce proteins. Nucleus m. RNA

The ribosome binds new t. RNA molecules and amino acids as it moves along the m. RNA. Phenylalanine Methionine Ribosome m. RNA Start codon t. RNA Lysine

Protein Synthesis

The process continues until the ribosome reaches a stop codon. Polypeptide Ribosome t. RNA m. RNA

Codon DNA Single strand of DNA Codon m. RNA Alanine Arginine Leucine Protein Amino acids within a polypeptide

Quiz 8 -3

The role of a master plan in a building is similar to the role of which molecule? a. DNA b. messenger RNA c. transfer RNA d. ribosomal RNA

A base that is present in RNA but NOT in DNA is a. thymine. b. cytosine. c. uracil. d. adenine.

The nucleic acid responsible for bringing individual amino acids to the ribosome is a. messenger RNA. b. DNA. c. transfer RNA. d. ribosomal RNA.

A region of a DNA molecule that indicates to an enzyme where to bind to make RNA is the a. intron. b. exon. c. codon. d. promoter.

A codon typically carries sufficient information to specify a(an) a. single base pair in RNA. b. single amino acid. c. entire protein. d. single base pair in DNA.

Mutations

Mutations are changes in the genetic material. Kinds of Mutations • Gene Mutations • Mutations that produce changes in a single gene. • Chromosomal Mutations • Mutations that produce changes in whole chromosomes.

Gene Mutations Gene mutations involving a change in one or a few nucleotides are known as point mutations because they occur at a single point in the DNA sequence. Point mutations include substitutions, insertions, and deletions. DNA Mutations Video

Substitutions usually affect no more than a single amino acid.

The effects of insertions or deletions are more dramatic. The addition or deletion of a nucleotide causes a shift in the grouping of codons. Changes like these are called frameshift mutations.

In an insertion, an extra base is inserted into a base sequence.

In a deletion, the loss of a single base is deleted and the reading frame is shifted.

Chromosomal Mutations • Chromosomal mutations involve changes in the number or structure of chromosomes. • Chromosomal mutations include deletions, duplications, inversions, and translocations.

Deletions involve the loss of all or part of a chromosome.

Duplications produce extra copies of parts of a chromosome.

Inversions reverse the direction of parts of chromosomes.

Translocations occurs when part of one chromosome breaks off and attaches to another.

Significance of Mutations • Many mutations have little or no effect on gene expression. • Some mutations are the cause of genetic disorders. • Polyploidy is the condition in which an organism has extra sets of chromosomes.

Polyploidy Down’s Syndrome Turner’s Syndrome

Quiz 8 -4

A mutation in which all or part of a chromosome is lost is called a(an) a. deletion. b. duplication. c. inversion. d. point mutation.

A mutation that affects every amino acid following an insertion or deletion is called a(an) a. point mutation. b. frameshift mutation. c. chromosomal mutation. d. inversion.

A mutation in which a segment of a chromosome is repeated is called a(an) a. deletion. b. inversion. c. duplication. d. point mutation.

The type of point mutation that usually affects only a single amino acid is called a. a deletion. b. a substitution. c. an insertion. d. a frameshift mutation.

When two different chromosomes exchange some of their material, the mutation is called a(an) a. inversion. b. deletion. c. substitution. d. translocation.

• Gene Regulation: An Example • E. coli provides an example of how gene expression can be regulated. • An operon is a group of genes that operate together. • In E. coli, these genes must be turned on so the bacterium can use lactose as food. • Therefore, they are called the lac operon.

How are lac genes turned off and on?

–The lac genes are turned off by repressors and turned on by the presence of lactose.

On one side of the operon's three genes are two regulatory regions. • In the promoter (P) region, RNA polymerase binds and then begins transcription.

The other region is the operator (O).

When the lac repressor binds to the O region, transcription is not possible.

• When lactose is added, sugar binds to the repressor proteins.

Gene Regulation: An changes shape and Example • The repressor protein falls off the operator and transcription is made possible.

• Many genes are regulated by repressor proteins. • Some genes use proteins that speed transcription. • Sometimes regulation occurs at the level of protein synthesis.

• trp Operon

• lac vs. trp Operons

How are most eukaryotic genes controlled?

• Eukaryotic Gene Regulation –Operons are generally not found in eukaryotes. –Most eukaryotic genes are controlled individually and have regulatory sequences that are much more complex than those of the lac operon.

• Many eukaryotic genes have a sequence called the TATA box. Upstream enhancer TATA box Promoter sequences Introns Exons Direction of transcription

• The TATA box seems to help position RNA polymerase. Upstream enhancer TATA box Promoter sequences Introns Exons Direction of transcription

• Eukaryotic promoters are usually found just before the TATA box, and consist of short DNA sequences. Upstream enhancer TATA box Promoter sequences Introns Exons Direction of transcription

• Genes are regulated in a variety of ways by enhancer sequences. • Many proteins can bind to different enhancer sequences. • Some DNA-binding proteins enhance transcription by: • opening up tightly packed chromatin • helping to attract RNA polymerase • blocking access to genes.

• Development and Differentiation • As cells grow and divide, they undergo differentiation, meaning they become specialized in structure and function. • Hox genes control the differentiation of cells and tissues in the embryo.

• Careful control of expression in hox genes is essential for normal development. • All hox genes are descended from the genes of common ancestors.

• Hox Genes Fruit fly chromosome Mouse chromosomes Fruit fly embryo Mouse embryo Adult fruit fly Adult mouse

Quiz 8 -5

–Which sequence shows the typical organization of a single gene site on a DNA strand? a. start codon, regulatory site, promoter, stop codon b. regulatory site, promoter, start codon, stop codon c. start codon, promoter, regulatory site, stop codon d. promoter, regulatory site, start codon, stop codon

– A group of genes that operates together is a(an) a. promoter. b. operon. c. operator. d. intron.

– Repressors function to a. turn genes off. b. produce lactose. c. turn genes on. d. slow cell division.

–Which of the following is unique to the regulation of eukaryotic genes? a. promoter sequences b. TATA box c. different start codons d. regulatory proteins

–Organs and tissues that develop in various parts of embryos are controlled by a. regulation sites. b. RNA polymerase. c. hox genes. d. DNA polymerase.