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
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
Related questions
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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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