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d) Cleavage by trypsin followed by gel-filtration chromatography gives the expected 6 products, which you sequence (shown in order of emergence from column): T-1 ETMESSAGEFGR T-2 SQTWALDHSECR T-4 TCR S-5 TME Think About It: T-5 NP e) Cleavage by Staph. aureus V8 protease followed by gel-filtration chromatography gives the expected 7 products, which you sequence (shown in order of emergence from column): S-1 RSQTWALD S-2 FGRGPQD S-3 NKTCRNP S-6 CRE T-3 GPQDNK S-7 HSE T-6 R S-4 SSAGE 1. Deduce the primary structure of this polypeptide. 2. Why would cyanogen bromide not be a good choice as a cleavage reagent? 3. Can you account for the order of elution of trypsin digest peptides from gel-filtration chromatography?

d) Cleavage by trypsin followed by gel-filtration chromatography gives the expected 6 products, which you sequence (shown in order of emergence from column): T-1 ETMESSAGEFGR T-2 SQTWALDHSECR T-4 TCR S-5 TME Think About It: T-5 NP e) Cleavage by Staph. aureus V8 protease followed by gel-filtration chromatography gives the expected 7 products, which you sequence (shown in order of emergence from column): S-1 RSQTWALD S-2 FGRGPQD S-3 NKTCRNP S-6 CRE T-3 GPQDNK S-7 HSE T-6 R S-4 SSAGE 1. Deduce the primary structure of this polypeptide. 2. Why would cyanogen bromide not be a good choice as a cleavage reagent? 3. Can you account for the order of elution of trypsin digest peptides from gel-filtration chromatography?

Biochemistry
Biochemistry
9th Edition
ISBN:9781319114671
Author:Lubert Stryer, Jeremy M. Berg, John L. Tymoczko, Gregory J. Gatto Jr.
Publisher:Lubert Stryer, Jeremy M. Berg, John L. Tymoczko, Gregory J. Gatto Jr.
Chapter1: Biochemistry: An Evolving Science
Section: Chapter Questions
Problem 1P
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**Educational Transcription: Analyzing Peptide Sequences** --- ### Peptide Analysis in South American Rainforest Research **Context:** In this scenario, researchers are exploring naturally occurring peptides from tropical orchids in a South American rainforest, aiming to identify those with antiviral potential. **Available Tools:** - Mobile biochemistry lab - Common reagents and enzymes - Amino-acid analyzer - Gel-filtration and ion-exchange chromatography - Electrophoresis - Edman Sequenator (limited by contaminants) **Objective:** Determine the amino acid sequence of a discovered peptide using available techniques. --- **Steps and Results:** **a) Molecular Weight Estimation:** - **Technique:** Electrophoresis - **Result:** MW ≈ 4000 **b) Amino Acid Analysis:** - **Technique:** Fragmentation assistance - **Result:** Identified sequence A\(_2\)C\(_2\)D\(_2\)E\(_4\)FG\(_3\)HKLMN\(_2\)P\(_2\)Q\(_2\)R\(_4\)S\(_4\)T\(_3\)W **c) Predicted Peptides via Cleavage Reagents:** - **Cyanogen Bromide (C-side of M)** - **Staphylococcus aureus V8 Protease (C-side of D and E)** - **Trypsin (C-side of K and R)** **d) Trypsin Cleavage and Gel-Filtration:** **Products Obtained:** - T-1: ETMESSAGEFGR - T-2: SQTWALDHSECR - T-3: GPQDNK - T-4: TCR - T-5: NP - T-6: R **e) Staph. aureus V8 Protease Cleavage:** **Products Obtained:** - S-1: RSQTWALD - S-2: FGRGPQD - S-3: NKTCNRNP - S-4: SSAGE - S-5: TME - S-6: CRE - S-7: HSE --- **Considerations and Problem Solving:** 1. **Deducing the Primary Polypeptide Structure
Transcribed Image Text:**Educational Transcription: Analyzing Peptide Sequences** --- ### Peptide Analysis in South American Rainforest Research **Context:** In this scenario, researchers are exploring naturally occurring peptides from tropical orchids in a South American rainforest, aiming to identify those with antiviral potential. **Available Tools:** - Mobile biochemistry lab - Common reagents and enzymes - Amino-acid analyzer - Gel-filtration and ion-exchange chromatography - Electrophoresis - Edman Sequenator (limited by contaminants) **Objective:** Determine the amino acid sequence of a discovered peptide using available techniques. --- **Steps and Results:** **a) Molecular Weight Estimation:** - **Technique:** Electrophoresis - **Result:** MW ≈ 4000 **b) Amino Acid Analysis:** - **Technique:** Fragmentation assistance - **Result:** Identified sequence A\(_2\)C\(_2\)D\(_2\)E\(_4\)FG\(_3\)HKLMN\(_2\)P\(_2\)Q\(_2\)R\(_4\)S\(_4\)T\(_3\)W **c) Predicted Peptides via Cleavage Reagents:** - **Cyanogen Bromide (C-side of M)** - **Staphylococcus aureus V8 Protease (C-side of D and E)** - **Trypsin (C-side of K and R)** **d) Trypsin Cleavage and Gel-Filtration:** **Products Obtained:** - T-1: ETMESSAGEFGR - T-2: SQTWALDHSECR - T-3: GPQDNK - T-4: TCR - T-5: NP - T-6: R **e) Staph. aureus V8 Protease Cleavage:** **Products Obtained:** - S-1: RSQTWALD - S-2: FGRGPQD - S-3: NKTCNRNP - S-4: SSAGE - S-5: TME - S-6: CRE - S-7: HSE --- **Considerations and Problem Solving:** 1. **Deducing the Primary Polypeptide Structure
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