From Gene to Protein

Important words and concepts from Chapter 17, Campbell & Reece, 2002 (1/29/2005):

by Stephen T. Abedon (abedon.1@osu.edu) for Biology 113 at the Ohio State University

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(1) Chapter title: From Gene to Protein

(a) “The DNA inherited by an organism leads to specific traits by dictating the synthesis of certain proteins. Proteins are the links between genotype and phenotype.”

(b) [from gene to protein (Google Search)] [index]

(2) Central dogma of molecular genetics (reverse transcription)

(a) The central dogma of molecular genetics is typically depicted as a shorthand review of how genetic information moves around a cell, or from parent to offspring.

(b) The central dogma looks like this:

(i) DNA ß DNA à RNA à protein

(i) DNA à DNA = replication (direction of arrow is arbitrary)

(ii) DNA à RNA = transcription (direction of arrow is not arbitrary)

(iii) RNA à protein = translation (ditto)

(d) This chapter deals particularly with the last two, transcription and translation

(e)

(f) (reverse transcription serves as an exception to the central dogma as originally conceived; it consists of DNA ß RNA, i.e., RNA à DNA, and is employed by such things as retroviruses including the virus that causes AIDS; note in the above figure that, of course, proteins also serve as enzymes)

(g) [central dogma, central dogma molecular genetics (Google Search)] [index]

(3) RNA (uracil)

(a) See Figure 17.2, Overview: the roles of transcription and translation in the flow of genetic information

(b) RNA is a nucleic acid polymer that resembles DNA except

(i) RNA uses the sugar ribose instead of deoxyribose

· Ribose has an –OH group at the 2’ carbon instead of the –H seen with deoxyribose found in DNA

(ii) RNA employs the nitrogenous base uracil (U) instead of the pyrimidine thymine

· For the latter point, that is, T » U

· The analogous base-pairing is U-A

· Note that U is energetically cheaper to make than T but that U is also less stable than T

(4) One gene-one polypeptide hypothesis

(a) Beadle and Tatum developed the one gene-one enzyme hypothesis in the 1940s

(b) The idea is that Mendel’s hereditary units are found in DNA but work by specifying enzymes

(c) This hypothesis was modified to one gene-one protein since not all proteins are enzymes but genes work by specifying proteins

(d) Finally, this hypothesis was modified to one gene-one polypeptide since many proteins consist of more than one polypeptide

(e) Genes specify the construction of specific polypeptides

(f) (in fact, to deconstruct things further, genes specify the transcription of specific RNAs)

(g) See Figure 17.1, Beadle and Tatum’s evidence for the one gene—one enzyme hypothesis

(h) ["one gene one protein", "one gene one polypeptide", "one gene one peptide" (Google Search)] [index]

TRANSCRIPTION

(5) Transcription—introduction (template strand)

(a) The DNA à RNA flow of genetic information is termed transcription

(b) The term transcription reflects that the information in DNA (i.e., nucleotide sequence) is copied into a similar code in RNA

(d) In different places on a chromosome the other strand may be copied

(e) The DNA strand that provides the complementary template to RNA polymerization is called the template strand

(f) The RNA can be of a number of types including:

(i) Messenger RNA (mRNA)

(ii) Transfer RNA (tRNA)

(iii) Ribosomal RNA (rRNA)

(iv) Etc. (e.g., spliceosomes)

(g) See Figure 17.2, Overview: the roles of transcription and translation in the flow of genetic information

(h) (see transcription in detail, below)

(i) [RNA transcription, template strand (Google Search)] [index]

(6) Messenger RNA (mRNA)

(a) If the RNA produced by transcription is to be used to code for the synthesis of proteins, it is called messenger RNA (a.k.a., mRNA)

(b) [messenger RNA, mRNA (Google Search)] [index]

(7) Transfer RNA (1) (tRNA)

(a) Another category of RNA, used during protein synthesis to ferry amino acids to growing peptide chains, is called transfer RNA (a.k.a., tRNA)

(b) (for more information, see transfer RNA, below)

(8) Ribosomal RNA (rRNA)

(a) Another category of RNA that together constitute about 60% of the mass of ribosomes is called ribosomal RNA or rRNA

(b) (in Escherichia coli cells, ribosomes make up 25% of the dry weight of cells)

(9) Translation—introduction

(a) The RNA à protein flow of genetic information is termed translation

(b) The term translation reflects that the information in mRNAs (i.e., nucleotide sequence) is translated into a new “language”, i.e., amino acid sequence

(c) See Figure 17.2, Overview: the roles of transcription and translation in the flow of genetic information

(d) (see translation in detail, below)

(e) [protein translation (Google Search)] [index]

(10) Eucaryotic segregation of transcription and translation

(a) Note that due to the existence of the nuclear membrane in eucaryotes, there exists a temporal and spatial separation of transcription and translation

(b) See Figure 17.2, Overview: the roles of transcription and translation in the flow of genetic information

(d) Translation occurs within the cytosol, where the functional ribosomes reside

(e) There is no such segregation of transcription and translation in prokaryotes

(f) [segregation of translation and transcription (Google Search)] [index]

(11) Codons

(a) The DNA and RNA nucleotide sequence code consists of one of four types of nucleotides (4 each, that is)

(b) The amino acid sequence code consists of 20 amino acids

(d) There also cannot be a two-to-one correspondence (4² < 20)

(e) Instead there exists a three to one correspondence (4³ > 20)

(f) The three nucleotides that specify an amino acid during translation are called codons

(g) See Figure 17.3, The triplet code

(h) See Figure 17.4, The dictionary of the genetic code

(i) [codons or codon (Google Search)] [the genetic code (the table of codons and what that means) (Shaun D. Black)] [index]

(12) Codons are a property of mRNA

(a) Note that codons exist in mRNA, but only their complement exists on the template strand of DNA

(b) (though note, additionally, that on the non-template strand of DNA the analogous DNA codons—though without uracil—exists)

(c) See Figure 17.4, The dictionary of the genetic code

(d) [codons mRNA (Google Search)] [index]

(13) Redundancy of triplet code

(a) 4³ = 64 >> 20

(b) Consequently, there are many more codons than there are amino acids

(d) This is because many amino acids are specified by more than one codon

(e) See Figure 17.4, The dictionary of the genetic code

(f) (no, you don’t have to memorize the figure)

(g) [triplet code redundancy OR redundant (Google Search)] [index]

(14) Lack of ambiguity in the triplet code

(a) Note that while the code is redundant, it is not ambiguous

(b) That is, each codon specifies for one and only one amino acid, not more than one

(15) Codons don’t overlap

(a) Another property of codons is that they are arrayed one after another in the mRNA

(b) That is, they do not overlap

(c) (note that there is an only slightly related exception in which codons can overlap and this is when reading frames of different genes overlap)

(d) See Figure 17.3, The triplet code

(e) [codons overlap (Google Search)] [index]

(16) There is no punctuation between codons

(a) Furthermore, codons do not have gaps between them (i.e., there is no punctuation)

(b) See Figure 17.3, The triplet code

(17) Start codon (AUG, methionine)

(a) The codon AUG codes for the amino acid methionine

(b) See Figure 17.4, The dictionary of the genetic code

(d) Thus, all polypeptides initially begin with methionine (Met)

(e) Note that as a part of post-translational protein processing the Met amino acid is often clipped off

(f) (though I don’t expect you to learn all of the codons and their assignments, you should memorize AUG, methionine, and the fact that it serves as the start codon of reading frames)

(g) [start codon, methionine (Google Search)] [index]

(18) Stop codons (nonsense codons)

(a) Only 61 of the 64 possible codons specify amino acids

(b) The other three specify what are known as stop codons

(d) See Figure 17.4, The dictionary of the genetic code

(e) Stop codons instruct the ribosome to stop adding amino acids to the growing peptide chain

(f) [stop codon (Google Search)] [index]

(19) Reading frame

(a) The sequence of codons beginning with AUG and ending with a stop codon is called the reading frame

(b) Note that the reading frame consists of (x + 1) * 3 nucleotides where x is the number of amino acids found in the resulting polypeptide (prior to post-translational modification) and the additional 1 is a stop codon

(20) (nearly) Universal triplet code

(a) The language of codons is nearly universal among extant organisms

(b) (e.g., AUG specifies Met and is the start codon in all or nearly all living organisms)

(d) Furthermore, the divergence from this common ancestor must have occurred at a time after the implementation of the triplet code

(e) Since the triplet code is somewhat arbitrary, the converse hypothesis, that all organisms somehow independently adopted the same codons for each amino acid, is much less likely

(f) As a consequence of the near-universality of the triplet code, genes from one organism may be transferred into unrelated organisms and still express (i.e., be transcribed then translated)

(g) [universal triplet code (Google Search)] [index]

(21) Transcription in detail

(a) Transcription takes place in three steps

(i) DNA binding and initiation

(ii) Elongation of the RNA strand

(iii) Termination of transcription

(b) The primary enzyme involved is called RNA polymerase

(c) See Figure 17.6, The stages of transcription: initiation, elongation, and termination

(d) See Figure 17.25, A summary of transcription and translation in a eukaryotic cell

(e) [RNA transcription (Google Search)] [index]

(22) RNA polymerase

(a) RNA polymerase works similarly to DNA polymerase

(b) Like DNA polymerase, RNA polymerase employs a DNA template (i.e., the template strand) but, of course, polymerizes RNA

(d) [RNA polymerase (Google Search)] [index]

(23) Promoter binding (transcription factor)

(a) The first step in transcription is DNA binding

(b) In prokaryotes this involves the recognition of specific DNA sequences (promoters) by the RNA polymerase

(c) See Figure 17.7, The initiation of transcription in a eukaryotic promoter

(d) In either case, the promoter is found upstream from the start codon

(e) Once bound the RNA polymerase begins transcribing (i.e., polymerizing RNA from a DNA template)

(f) In eukaryotes this involves the binding of RNA polymerase to proteins, called transcription factors, that are involved in sequence recognition

(g) [promoter binding (Google Search)] [index]

(24) Elongation (1)

(a) To initiate transcription, the RNA polymerase must separate the DNA strands of the double helix

(b) Throughout the elongation of the RNA transcript, the DNA strand is kept open approximately 10 bases

(c) Note that a given gene may be transcribed by more than one RNA polymerase simultaneously, with one RNA polymerase following another along on the transcribed DNA

(d) See Figure 17.6, The stages of transcription: initiation, elongation, and termination

(e) [transcription elongation (Google Search)] [index]

(25) Termination of transcription

(a) Just as transcription is initiated at certain base sequences, it is similarly terminated at specific base sequences

(b) With termination the RNA transcript is released from the RNA polymerase and DNA template strand, and the RNA polymerase from the DNA

(c) See Figure 17.6, The stages of transcription: initiation, elongation, and termination

(d) [transcription termination (Google Search)] [index]

(26) mRNA processing

(a) The compartmentalization of the eukaryotic cell results in a separation of transcription and translation, both spatially and temporally

(b) Eukaryotic cells take advantage of this compartmentalization to modify RNAs prior to translation

(i) Addition of a 5’ cap

(ii) Addition of a poly-A tail

(iii) Removal of introns

(d) See Figure 17.8, RNA processing: addition of 5’ cap and poly(A) tail

(e) mRNAs are allowed to leave the nucleus only once they have been processed

(f) (recall that translation occurs only in the cytosol)

(g) [mRNA processing (Google Search)] [index]

(27) Introns and exons

(a) Most eukaryotic genes do not exist as continuous reading frames

(b) Eukaryotic mRNAs, however, do exist as continuous reading frames

(d) The intervening sequences that disrupt reading frames in genes and in RNAs prior to their processing are called introns (i.e., intervening sequences)

(e) The sequences which are spliced together, upon the removal of introns, to form a continuous reading frame are called exons (i.e., expressed sequences)

(f) See Figure 17.9, RNA processing: RNA splicing

(g) [introns exons (Google Search)] [index]

(28) Spliceosome

(a) There exists a nuclear structure involved in intron excision called a spliceosome

(b) Note that yet another form of RNA plays a functional role in spliceosomes

(c) See Figure 17.10, The roles of snRNPs and splicosomes in mRNA splicing

(d) [spliceosome (Google Search)] [index]

TRANSLATION

(29) Translation in detail

(a) Translation is far more complex than transcription, involving many dozens of distinct macromolecular players

(b) See Figure 17.12, Translation, the basic concept

(i) Transfer RNAs

(ii) Ribosomes

(iii) Aminoacyl-tRNA-synthetases

(iv) mRNA

(v) The growing peptide

(d) Analogously, transcription employs DNA, RNA polymerase, and a growing RNA transcript

(e) Like transcription, translation occurs in three basic steps

(i) Initiation

(ii) Elongation

(iii) Termination

(f) Note that much of translation is powered by the nucleoside triphosphate GTP (which you last saw during the Krebs cycle) rather than ATP

(g) Remember that the primary goal of translation is the synthesis of a polypeptide from mRNA-coded information

(h) Also, keep in mind that it is probably much easier to understand translation by following the figures in your text than from simply reading the text or these lecture notes

(i) See Figure 17.23, A summary of transcription and translation in a eukaryotic cell

(j) [translation RNA (Google Search)] [index]

(30) Transfer RNA (2)

(a) Transfer RNAs are the translating units

(b) One side of the tRNA binds to a specific codon found on an mRNA

(d) See Figure 17.12, Translation, the basic concept

The Genetic Code (supplemental table)
	U	C	A	G
U	Phenylalanine	Serine	Tyrosine	Cysteine	U
					C
	Leucine		STOP	STOP	A
				Tryptophan	G
C	Leucine	Proline	Histidine	Arginine	U
					C
			Glutamine		A
					G
A	Isoleucine	Threonine	Asparagine	Serine	U
					C
			Lysine	Arginine	A
	Methionine				G
G	Valine	Alanine	Aspartic Acid	Glycine	U
					C
			Glutamic Acid		A
					G

(e) See Figure 17.13, The structure of transfer RNA (tRNA)

(f) [transfer RNA, tRNA (Google Search)] [index]

(31) Anticodon

(a) The region of the tRNA that binds to the mRNA codon is called the anticodon

(b) See Figure 17.12, Translation, the basic concept

(d) [anticodon (Google Search)] [index]

(32) Wobble

(a) This complementarity between codon and anticodon is “more or less” because the third base of a codon tends to be ambiguously bound by the anticodon

(b) Note that much of the variation in the sequence of the codons specifying individual amino acids is found in the third base of the codon

(d) This tendency of anticodons to bind codons varying in their third base is called wobble (see table to right)

(e) See Figure 17.4, The dictionary of the genetic code (to see how the third bases in codons tend to have a more-minor role in specifying the amino acid than the first two bases)

(f) See Figure 17.13, The structure of transfer RNA (tRNA) (note that it is the 5’ base that is read ambiguously due to wobble)

(g) [wobble translation (Google Search)] [index]

(33) Aminoacyl-tRNA synthetases

(a) tRNAs employ their anticodons to bind specific codons found on mRNAs

(b) However, tRNAs are not responsible for specifying what amino acid they attach to

(d) These enzymes are called aminoacyl-tRNA-synthetases

(e) (yes, it is a big word; sound it out as: amino-acyl-tRNA-syn-theh-tase)

(f) The cost of amino acid addition is one ATP

(g) See Figure 17.14, An aminoacyl-tRNA synthetase joins a specific amino acid to a tRNA

(h) There exist at least one aminoacyl-tRNA-synthetase for each amino acid (i.e., 20)

(i) [aminoacyl-tRNA synthetase (Google Search)] [index]

(34) Ribosomes (A site, P site, E site)

(a) Ribosomes are the machines within which tRNAs function to read the mRNA code and translate that code into polypeptides

(b) Ribosomes consist of one large and one small subunit (both which are complexes of rRNA and many proteins)

(i) The mRNA

(ii) The tRNA attached to the incoming amino acids (the A site)

(iii) The tRNA attached to the growing polypeptide (the P site)

(d) Ribosomes additionally have an “E site” from which tRNAs exit the ribosome

(e) See Figure 17.15, The anatomy of a functioning ribosome

(f) The ribosome’s function is to catalyze the peptide bond formation between the polypeptide held at the P site to the incoming amino acid held at the A site

(g) [ribosome (Google Search)] [index]

(35) Initiation

(a) Initiation of translation involves the binding of the Met-carrying tRNA to the AUG start codon found on the mRNA that in turn is bound to the small subunit of the ribosome

(b) See Figure 17.17, The initiation of translation

(d) Note that the tRNA is found at what will be the P site

(e) The large ribosomal subunit then binds to the small subunit

(f) The above binding occurs at a cost of one GTP

(g) [translation elongation (Google Search)] [index]

(36) Elongation (2)

(a) Elongation in translation is more complex than that of transcription because there are more players (e.g., tRNAs and ribosomes) and because the mRNA is read three nucleotides (one codon) at a time rather than only a single nucleotide (i.e., as in transcription)

(b) Charged tRNAs (i.e., ones to which an amino acid is bound) diffuse into the A site and only those that successfully interact with the mRNA codon stay there (this step actually requires energy to perform—one GTP)

(c) A peptide bond is then formed between the incoming amino acid and the peptide held at the P site (this releases the peptide from the P-site located tRNA—no additional energy is required from that already stored in the various molecules involved)

(d) The mRNA is then translocated one codon forward so that the tRNA that had held only one amino acid in the A site, but now holds the growing polypeptide, is now found in the P site

(e) The translocation step requires one GTP

(f) See Figure 17.18, The elongation cycle of translation

(g) Note that the ribosome moves along the mRNA in the 5’ à 3’ direction—the mRNA thus moves through the ribosome in the 3’ à 5’ direction (i.e., the 5’ end leads)

(h) [translation elongation (Google Search)] [index]

(37) Termination

(a) When the codon in the A site is a nonsense (stop) codon, no associated tRNA exists

(b) Instead release factors bind to the stop codon in the A site

(d) In addition, the ribosome releases the mRNA and then separates into two subunits

(e) See Figure 17.19, The termination of translation

(f) [translation termination (Google Search)] [index]

(38) Post-translational polypeptide modification

(a) “During and after its synthesis, a polypeptide chain begins to coil and fold spontaneously, forming a functional protein of specific conformation: A three-dimensional molecule with secondary and tertiary structures. A gene determines primary structure, and primary structure determines conformation.”

(b) Posttranslational modifications of this folding polypeptide, however, can include

(i) Covalent attachment of sugars, lipids, phosphate groups, etc.

(ii) Removal of one or more leading (amino) end amino acids (e.g., Met)

(iii) Cleavage of polypeptide chain

(39) Signal sequences

(a) The targeting of proteins occurs during and after translation

(b) An amino-terminal amino acid sequence can play a role in protein targeting

(d) “Signal sequences function like ZIP codes, addressing proteins to certain locations in the cell.”

(e) See Figure 17.21, The signal mechanisms for targeting proteins to the ER

(f) [signal sequence (Google Search)] [index]

MUTATION

(40) Mutation

(a) A mutation is a replicable change in nucleotide sequence

(b) Contrast this with DNA damage which is a non-replicable alteration in DNA structure

(i) Point mutations

(ii) Silent mutations

(iii) Missense mutations

(iv) Nonsense mutations

(v) Insertions

(vi) Deletions

(vii) Frameshift mutations

(d) [mutation (Google Search)] [index]

(41) Mutations are typically detrimental . . .

(a) Mutations represent a change in highly evolved information

(b) Typically changes to well functioning systems are detrimental, and muta tions are no exception

(d) [mutations detrimental, mutations bad (Google Search)] [index]

(42) …but Mutations are the only way to change

(a) Despite the typically detrimental nature of mutations, they also represent the only way in which novel, beneficial information is typically introduced into a genetic system

(b) (the other way is horizontal transfer, i.e., from different species, and even then mutations still represent the ultimate source of information)

(c) Thus, while mutations typically are detrimental, from time to time a mutation actually increases the survival and reproductive potential of an organism

(43) Point mutation

(a) Mutations come in a number of “flavors”, the easiest to envisage being the point mutation

(b) A point mutation is simply a change of one nucleotide in a base sequence to a different nucleotide (e.g., a change from A to G)

(c) See Figure 17.23, The molecular basis of sickle-cell disease: a point mutation

(d) Upon replication that change will be duplicated into the complementary strand of one of the daughter chromosomes

(e) [point mutation (Google Search)] [index]

(44) Silent mutation

(a) Point mutations, even those occurring within exons , need not result in changes to amino-acid sequence

(b) Why? Recall that third base substitutions in codons frequently will not result in a change in the amino acid coded for

(45) Missense mutation

(a) When a point mutation results in a change in amino acid, that mutation is termed a missense mutation

(b) See Figure 17.24, Categories and consequences of point mutations

(d) If the mutation occurs in a less crucial region of a polypeptide, or results in a change to a functionally similar amino acid, no significant impact may occur

(e) If the mutation occurs in the active site or other crucial region of the polypeptide, significant impact may occur

(f) Note that typically any impact will be detrimental

(g) [missense mutation (Google Search)] [index]

(46) Nonsense mutation

(a) A nonsense mutation is a point mutation that results in a change from an amino acid-coding codon (a.k.a., a sense codon) to a stop codon (a.k.a., nonsense codon)

(b) See Figure 17.24, Categories and consequences of point mutations

(d) Note also that not all sense codons can be converted to a nonsense codon via only a single point mutation

(e) [nonsense mutation (Google Search)] [index]

(47) Insertion

(a) An insertion increases the number of nucleotides in a sequence

(b) [insertion mutation (Google Search)] [index]

(48) Deletion

(a) A deletion decreases the number of nucleotides in a sequence

(b) [deletion mutation (Google Search)] [index]

(49) Frameshift mutation

(a) Insertions or deletions of more or less than multiples of three cause the most significant disruption