Podcast
Questions and Answers
Match the following reagents with their specific cleaving actions:
Match the following reagents with their specific cleaving actions:
Chymotrypsin = Hydrolysis of peptide bonds on the carboxyl side of Lys and Arg Trypsin = Cleavage of peptide bonds on the carboxyl side of Met Dithiothreitol = Breakage of disulfide (—S—S—) bonds Edman reagent (phenylisothiocyanate) = Determination of the amino acid sequence of a peptide FDNB = Determining the amino-terminal amino acid in a polypeptide
What is a major step in determining the sequence of amino acid residues in a protein?
What is a major step in determining the sequence of amino acid residues in a protein?
Use Edman degradation to determine the sequence of each peptide.
What is the likely explanation for the identification of Ala and Leu as amino-terminal residues in equal amounts?
What is the likely explanation for the identification of Ala and Leu as amino-terminal residues in equal amounts?
The protein has some multiple of two subunits, with Ala and Leu as the amino-terminal residues.
What conclusion can be drawn about the protein that has undergone treatment with performic acid?
What conclusion can be drawn about the protein that has undergone treatment with performic acid?
Describe a key difference between chemical synthesis of polypeptides and synthesis in living cells.
Describe a key difference between chemical synthesis of polypeptides and synthesis in living cells.
What is the distinction between homologs, paralogs, and orthologs?
What is the distinction between homologs, paralogs, and orthologs?
How are 'signature sequences' useful in analyzing proteins?
How are 'signature sequences' useful in analyzing proteins?
Which two amino acids differ from each other by only one atom?
Which two amino acids differ from each other by only one atom?
Identify the pair of peptides below that cannot be distinguished by tandem mass spectrometry.
Identify the pair of peptides below that cannot be distinguished by tandem mass spectrometry.
List three classes of conjugated proteins and their associated prosthetic groups.
List three classes of conjugated proteins and their associated prosthetic groups.
What is a polymorphic protein?
What is a polymorphic protein?
What is the native sequence determined from the tetrapeptide composed of (Lys) 2, Phe, and Gly?
What is the native sequence determined from the tetrapeptide composed of (Lys) 2, Phe, and Gly?
When a gene is available and its sequence of nucleotides is known, what chemical studies are still necessary?
When a gene is available and its sequence of nucleotides is known, what chemical studies are still necessary?
What does the term 'proteome' refer to?
What does the term 'proteome' refer to?
What major problem did advances in mass spectrometry overcome?
What major problem did advances in mass spectrometry overcome?
Which tripeptides from the following options have the largest number of nonpolar R groups?
Which tripeptides from the following options have the largest number of nonpolar R groups?
The amino acid ______ has no free alpha-amino group.
The amino acid ______ has no free alpha-amino group.
What are the structural characteristics common to all amino acids found in naturally occurring proteins?
What are the structural characteristics common to all amino acids found in naturally occurring proteins?
Only one of the common amino acids has no free alpha-amino group.
Only one of the common amino acids has no free alpha-amino group.
Which tripeptide will yield DNP-tyrosine when reacted with l-fluoro-2,4-dinitrobenzene and hydrolyzed in acid?
Which tripeptide will yield DNP-tyrosine when reacted with l-fluoro-2,4-dinitrobenzene and hydrolyzed in acid?
What is the pI, and how is it determined for amino acids that have nonionizable R groups?
What is the pI, and how is it determined for amino acids that have nonionizable R groups?
What factors affect the elution order of molecules during size-exclusion chromatography?
What factors affect the elution order of molecules during size-exclusion chromatography?
Define the primary structure of a protein.
Define the primary structure of a protein.
Which reagents are used in protein chemistry?
Which reagents are used in protein chemistry?
The chirality of an amino acid results from the fact that its alpha carbon:
The chirality of an amino acid results from the fact that its alpha carbon:
Of the 20 standard amino acids, only ___________ is not optically active. The reason is that its side chain ___________.
Of the 20 standard amino acids, only ___________ is not optically active. The reason is that its side chain ___________.
Two amino acids of the standard 20 contain sulfur atoms. They are:
Two amino acids of the standard 20 contain sulfur atoms. They are:
All of the amino acids that are found in proteins, except for proline, contain a(n):
All of the amino acids that are found in proteins, except for proline, contain a(n):
Which of the following statements about aromatic amino acids is correct?
Which of the following statements about aromatic amino acids is correct?
Which of the following statements about cystine is correct?
Which of the following statements about cystine is correct?
The uncommon amino acid selenocysteine has an R group with the structure —CH2—SeH (pKa ~5). In an aqueous solution, pH = 7.0, selenocysteine would:
The uncommon amino acid selenocysteine has an R group with the structure —CH2—SeH (pKa ~5). In an aqueous solution, pH = 7.0, selenocysteine would:
Amino acids are ampholytes because they can function as either a(n):
Amino acids are ampholytes because they can function as either a(n):
Titration of valine by a strong base, for example NaOH, reveals two pK's. The titration reaction occurring at pK2 (pK2 = 9.62) is:
Titration of valine by a strong base, for example NaOH, reveals two pK's. The titration reaction occurring at pK2 (pK2 = 9.62) is:
In a highly basic solution, pH = 13, the dominant form of glycine is:
In a highly basic solution, pH = 13, the dominant form of glycine is:
For amino acids with neutral R groups, at any pH below the pI of the amino acid, the population of amino acids in solution will have:
For amino acids with neutral R groups, at any pH below the pI of the amino acid, the population of amino acids in solution will have:
At pH 7.0, converting a glutamic acid to gamma-carboxyglutamate, will have what effect on the overall charge of the protein containing it?
At pH 7.0, converting a glutamic acid to gamma-carboxyglutamate, will have what effect on the overall charge of the protein containing it?
At pH 7.0, converting a proline to hydroxyproline, will have what effect on the overall charge of the protein containing it?
At pH 7.0, converting a proline to hydroxyproline, will have what effect on the overall charge of the protein containing it?
What is the approximate charge difference between glutamic acid and alpha-ketoglutarate at pH 9.5?
What is the approximate charge difference between glutamic acid and alpha-ketoglutarate at pH 9.5?
The formation of a peptide bond between two amino acids is an example of a(n) ______________ reaction.
The formation of a peptide bond between two amino acids is an example of a(n) ______________ reaction.
The peptide alanylglutamylglycylalanylleucine has:
The peptide alanylglutamylglycylalanylleucine has:
An octapeptide composed of four repeating glycylalanyl units has:
An octapeptide composed of four repeating glycylalanyl units has:
At the isoelectric pH of a tetrapeptide:
At the isoelectric pH of a tetrapeptide:
Which of the following is correct with respect to the amino acid composition of proteins?
Which of the following is correct with respect to the amino acid composition of proteins?
The average molecular weight of the 20 standard amino acids is 138, but biochemists use 110 when estimating the number of amino acids in a protein of known molecular weight. Why?
The average molecular weight of the 20 standard amino acids is 138, but biochemists use 110 when estimating the number of amino acids in a protein of known molecular weight. Why?
In a conjugated protein, a prosthetic group is:
In a conjugated protein, a prosthetic group is:
Prosthetic groups in the class of proteins known as glycoproteins are composed of:
Prosthetic groups in the class of proteins known as glycoproteins are composed of:
For the study of a protein in detail, an effort is usually made to first:
For the study of a protein in detail, an effort is usually made to first:
In a mixture of the five proteins listed below, which should elute second in size-exclusion (gel-filtration) chromatography?
In a mixture of the five proteins listed below, which should elute second in size-exclusion (gel-filtration) chromatography?
By adding SDS (sodium dodecyl sulfate) during the electrophoresis of proteins, it is possible to:
By adding SDS (sodium dodecyl sulfate) during the electrophoresis of proteins, it is possible to:
To determine the isoelectric point of a protein, first establish that a gel:
To determine the isoelectric point of a protein, first establish that a gel:
The first step in two-dimensional gel electrophoresis generates a series of protein bands by isoelectric focusing. In a second step, a strip of this gel is turned 90 degrees, placed on another gel containing SDS, and electric current is again applied. In this second step:
The first step in two-dimensional gel electrophoresis generates a series of protein bands by isoelectric focusing. In a second step, a strip of this gel is turned 90 degrees, placed on another gel containing SDS, and electric current is again applied. In this second step:
The term specific activity differs from the term activity in that specific activity:
The term specific activity differs from the term activity in that specific activity:
Which of the following refers to particularly stable arrangements of amino acid residues in a protein that give rise to recurring patterns?
Which of the following refers to particularly stable arrangements of amino acid residues in a protein that give rise to recurring patterns?
Which of the following describes the overall three-dimensional folding of a polypeptide?
Which of the following describes the overall three-dimensional folding of a polypeptide?
The functional differences, as well as differences in three-dimensional structures, between two different enzymes from E.coli result directly from their different:
The functional differences, as well as differences in three-dimensional structures, between two different enzymes from E.coli result directly from their different:
One method used to prevent disulfide bond interference with protein sequencing procedures is:
One method used to prevent disulfide bond interference with protein sequencing procedures is:
A nonapeptide was determined to have the following amino acid composition: (Lys)2, (Gly) 2, (Phe) 2, His, Leu, Met. What are the results of the peptide's analysis with 1-fluoro-2,4-dinitrobenzene?
A nonapeptide was determined to have the following amino acid composition: (Lys)2, (Gly) 2, (Phe) 2, His, Leu, Met. What are the results of the peptide's analysis with 1-fluoro-2,4-dinitrobenzene?
Flashcards
Chirality in Amino Acids
Chirality in Amino Acids
Amino acids are chiral because the alpha carbon is bonded to four different groups.
Glycine's Chirality
Glycine's Chirality
Glycine is achiral because it has two hydrogens attached to the alpha carbon.
Sulfur-Containing Amino Acids
Sulfur-Containing Amino Acids
Methionine and cysteine are the only standard amino acids with sulfur.
Amino Group Presence
Amino Group Presence
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Aromatic Amino Acids
Aromatic Amino Acids
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Cystine Formation
Cystine Formation
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Amino Acid as Zwitterion
Amino Acid as Zwitterion
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Ampholyte
Ampholyte
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Valine Titration
Valine Titration
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Glycine in Basic Solution
Glycine in Basic Solution
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Isoelectric Point (pI)
Isoelectric Point (pI)
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Peptide Bond Formation
Peptide Bond Formation
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Peptide Bond Count
Peptide Bond Count
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Protein Purification
Protein Purification
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Isoelectric Point Determination
Isoelectric Point Determination
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Secondary Protein Structure
Secondary Protein Structure
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Tertiary Protein Structure
Tertiary Protein Structure
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Mass Spectrometry Advancements
Mass Spectrometry Advancements
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Protein Sequence Elucidation
Protein Sequence Elucidation
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Proteome Definition
Proteome Definition
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Protein Sequence Homology
Protein Sequence Homology
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Amino Acid Structure
Amino Acid Structure
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Uncommon Amino Acids
Uncommon Amino Acids
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Zwitterion Formation in Water
Zwitterion Formation in Water
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Study Notes
Chirality and Amino Acids
- Chirality in amino acids arises from the alpha carbon being bonded to four different groups.
- Glycine is the only standard amino acid that is not optically active due to its hydrogen side chain.
Sulfur-Containing Amino Acids
- Methionine and cysteine are the only standard amino acids that contain sulfur.
Amino Acids in Proteins
- All standard amino acids, apart from proline, contain an amino group.
- Aromatic amino acids absorb ultraviolet light, with tryptophan being more efficient than tyrosine.
Cystine and Disulfide Bonds
- Cystine is formed when the thiol group of cysteine is oxidized, resulting in a disulfide bridge.
Zwitterion Form in Solutions
- At pH 7.0, selenocysteine exists as a fully ionized zwitterion.
- Amino acids function as ampholytes, acting as both acids and bases.
Titration and pKa Values
- The titration curve for valine reveals two pKa values; at pK2, the reaction involves the deprotonation of the amino group.
- In a basic solution, glycine predominantly exists as a protonated species.
Charge Properties of Amino Acids
- Below the isoelectric point (pI), amino acids with neutral side chains carry a net positive charge.
- Gamma-carboxyglutamate conversion at pH 7.0 makes proteins more negatively charged.
Peptide Bonds and Structure
- Peptide bond formation is a condensation reaction resulting in the release of water.
- The number of peptide bonds in a peptide is one less than the number of amino acids.
Protein Purification and Analysis
- Protein purification is a crucial first step in detailed protein study.
- The isoelectric point of a protein can be determined by creating a stable pH gradient in the gel.
Protein Structural Levels
- Secondary structure refers to stable arrangements of amino acid residues, such as alpha-helices and beta-sheets.
- Tertiary structure encompasses the overall three-dimensional folding of a polypeptide.
Mass Spectrometry and Protein Studies
- Advances in mass spectrometry help address challenges related to analyzing macromolecules.
Identifying Protein Sequences
- Techniques like trypsin digestion and cyanogen bromide help elucidate amino acid sequences in peptides.
- The native sequence of peptides can be determined using HPLC and other biochemical methods.
Proteomics
- "Proteome" refers to the full complement of proteins encoded by an organism's DNA, highlighting its complexity and functional diversity.### Mass Spectrometry and Proteins
- Mass spectrometry advancements enable the analysis of macromolecules but face challenges related to gas-phase molecules.
Protein Sequence Comparison
- Protein sequences from four different proteins analyzed for evolutionary relationships.
- Evolutionary divergence is highlighted between proteins based on amino acid sequences.
- Specific relationships include:
- Protein 4 exhibiting the greatest evolutionary divergence in column B.
- Proteins 1 and 2 have notably diverged evolutionarily.
- Protein 4 shows significant homology to protein 1.
Amino Acid Structure
- All naturally occurring amino acids share a structure of an alpha carbon connected to:
- A carboxylic acid
- An amine group
- A hydrogen atom
- A variable side chain.
- Amino acids exist predominantly in the L configuration.
Uncommon Amino Acids
- Examples include 4-hydroxyproline, 5-hydroxylysine, and selenocysteine.
- These amino acids typically arise from post-translational modifications of standard amino acids.
Zwitterion Formation
- Amino acids become zwitterions in water because:
- Carboxylic acid groups ionize to form COO⁻.
- Amino groups become NH₃⁺ by protonation.
Titration and Buffering
- At pH 9.5, the NH₃⁺ group in amino acids is titrated to NH₂.
- The positively charged –NH₃⁺ stabilizes the negatively charged COO⁻, lowering its pKa.
- The pH region with the greatest buffering capacity is around the amino acid's pKa, indicated by a leveling off in titration curves.
Protein Purification and Analysis
- Specific activity of an enzyme is calculated as activity per protein concentration.
- Peptide bonds are stable due to high activation energy for hydrolysis.
- Techniques like ion-exchange and size-exclusion chromatography separate proteins based on charge and size, respectively.
Isoelectric Point (pI)
- The isoelectric point is reached at a pH where a molecule exhibits no net charge.
- Calculated as the average of two pKa values for amino acids without ionizable side chains.
Protein Structure Determination
- Steps for deducing the sequence of an amino acid chain post-disulfide bond reduction:
- Edman degradation for each peptide.
- Use proteases like trypsin for additional peptide generation.
- Overlap peptides to reconstruct the full sequence.
Protein Relationships
- Distinction between homologs (related proteins within a family), paralogs (duplicate genes within a species), and orthologs (genes in different species).
- Signature sequences are instrumental in linking functionally related proteins, enhancing evolutionary and functional analysis.
Chemical vs. Biological Synthesis
- Chemical synthesis is from carboxyl to amino terminus, while biological synthesis is the reverse.
- Cells can synthesize larger and more complex proteins compared to current chemical methods, which are limited to shorter polypeptides.### Amino Acid Differences
- Serine (Ser) and Cysteine (Cys) differ by only one atom.
- Importance of identifying variations at the atomic level in amino acid sequences.
Tandem Mass Spectrometry
- Certain peptides cannot be distinguished using tandem mass spectrometry due to similarities in their sequences.
- Example pair: VTSPLYANEGK and VTSPIYANEGK are indistinguishable.
Conjugated Proteins
- Conjugated proteins contain additional chemical substituents beyond amino acids.
- Three classes of conjugated proteins include:
- Lipoproteins, associated with lipid groups
- Glycoproteins, associated with carbohydrate groups
- Phosphoproteins, associated with phosphoryl groups
- Hemoproteins, associated with heme groups
- Flavoproteins, associated with flavin nucleotide groups
- Metalloproteins, associated with metal ions (e.g., zinc, iron, calcium)
Polymorphic Proteins
- Polymorphic proteins are characterized by varying amino acid sequences among individuals in the human population.
- Variants of polymorphic proteins typically maintain similar function and activity.
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Description
Test your knowledge on various reagents used in the hydrolysis and cleavage of peptide bonds. This quiz covers important reagents like CNBr, Edman reagent, and enzymes such as chymotrypsin and trypsin, as well as their specific applications. Assess your understanding of protein chemistry and amino acid sequence determination.
