I want to separate a protein from a crude yeast extract by ion exchange at pH...
I want to separate a protein from a crude yeast extract by ion exchange at pH 7.4. Should I use a DEAE column or CM cellulose column? The isoelectric point (pI) of the protein is greater than the pH (pI = 9). Explain.
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I want to separate a protein from a crude yeast extract by ion
exchange at pH...
NOTES: To separate on a size-exclusion column, differences in MW need to be greater than 10%....
NOTES: To separate on a size-exclusion column, differences in MW need to be greater than 10%. To separate proteins by charge, the difference in pI must be at least 0.5 pH unit. Consider the table of four proteins below: Protein Size pI Size exclusion Charge at pH 7.4 DEAE-elution 1 49 kDa 6.8 2 51 kDa 8.1 3 53 kDa 6.4 4 99 kDa 8.3 5 106 kDa 3.8 6 143 kDa 7.9 (a) In what order would the proteins...
SIRJE GEL OBJECTIVE: Partial purification of a protein, isocitrate dehydrogenase (IDH), using DEAE ion exchange chromatography....
SIRJE GEL OBJECTIVE: Partial purification of a protein, isocitrate dehydrogenase (IDH), using DEAE ion exchange chromatography. IDH will be eluted from the column using increasing concentrations of KCI. IDH will be tracked using the IDH activity assay and the extent of purification will be analyzed using SDS PAGE Last week we assayed for the activity of the enzyme, isocitrate dehydrogenase, from E.coli strains carrying a plasmid that over-expresses the E. coli IDH gene. To purify proteins from cel extracts, biochemists...
______ 2. Using the accompanying table, determine which protein would stick to a carboxymethyl cellulose ion...
______ 2. Using the accompanying table, determine which protein would stick to a carboxymethyl cellulose ion exchange column (the column has stationary carboxyl groups) at pH 7.0. a) glucose oxidase b) horse myoglobin c) ribonuclease A d) cytochrome C Protein Isoelectric Point Amyloglucosidase Glucose oxidase Trypsin inhibitor β-Lactoglobulin Carboanhydrase Myoglobin horse Lentil lectin Ribonuclease A Cytochrome C pI 3.5 pI 4.2 pI 4.5 pI 5.15/5.3 pI 6.0 pI 7.35 pI 7.75/8.0/8.3 pI 9.45 pI 10.65
In your laboratory, you plan to use ion-exchange chromatography to separate the peptide shown below from...
In your laboratory, you plan to use ion-exchange chromatography to separate the peptide shown below from a mixture of different peptides at pH 7.0. Should you choose a matrix containing DEAE (+ charge) or CM (- charge) groups? GLEKSLVRLGDVQPSLGKESRAKKFQRQ
In order to separate protein X using ion exchange chromatography, I am using a resin with...
In order to separate protein X using ion exchange chromatography, I am using a resin with carboxylates attached (pka is 6.0). If I wanted to separate protein X, whose plis 8.0, what is the pH range of the buffer that can be used for ion exchange chromatography with this resin? any pH between 6.0 and 8.0 any pH above 6.0. any pH below 8.0 there is no pH range for this resin and protein X. Which of the following is...
Let's say that an unknown protein has an isoelectric point of 7.0. a) Will this protein...
Let's say that an unknown protein has an isoelectric point of 7.0. a) Will this protein be positively or negatively charged at pH 4.0? b) Will this protein be an anion or a cation at pH 10.0? c) If you had at your disposal a cation exchange column, would you separate the protein from others in the mix on the basis of cation exchange or anion exchange? Explain. d) Would phosphate buffer (pKa 7.20), ethanolamine buffer (pKa 9.50), or citric...
a-d plz 7. Protein purification. As a graduate student, the first protein I purified was RNA...
a-d plz
7. Protein purification. As a graduate student, the first protein I purified was RNA polymerase (RNAP) from E. coli- Some physical and chemical properties of Ecoli RNAP Molecular max470,000 g/mol polynentide compasitian (subunits): (50 kDa), k andok a pl = 5.34 substrates: NTPs cofactor: Mg? Purification protocol. E coli cells were broken usine Isozyme, yielding a cellualar extract containing a protcome solution 4M (NHOSO, was added to the cellular extract. A white protein precipitate was formed incuding RNAM...
7. Protein purification. As a graduate student, the first protein I purified was RNA polymerase (RNAP)...
7. Protein purification. As a graduate student, the first protein I purified was RNA polymerase (RNAP) from E. coli. Some physical and chemical properties of E. coli RNAP Molecular mass = 470,000 g/mol polypeptide composition (subunits): a (50 kDa), B (150 kDa) and 6 (70 kDa) pl = 5.34 substrates: NTPS cofactor: Mg Purification protocol. E. coli cells were broken using lysozyme, yielding a cellualar extract containing a proteome solution. 4M (NH4)2SO4 was added to the cellular extract. A white...
4. (Challenge) You are purifying a protein from the brain of patients with an unusual type of...
4. (Challenge) You are purifying a protein from the brain of patients with an unusual type of dementia triggered by a coronavirus and you hypothesize that this protein may be a signature molecule of this type of dementia. To pursue this hypothesis, you collect brain tissue from patients post mortem, do differential centrifugation, density gradient centrifugation, CM (carboxymethyl cellulose) ion exchange chromatography and gel filtration. You then assess the purity of your protein using both native and SDS gel electrophoresis. The...
1. A biochemist is attempting to separate a DNA-binding protein (protein X) from other proteins in...
1. A biochemist is attempting to separate a DNA-binding protein (protein X) from other proteins in a solution. Only three other proteins (A, B, and C) are present. The proteins have the following properties: pl (isoelectric point) Size Mr Bind to DNA? protein A 7.4 protein B 3.8 protein C 7.9 protein X7.8 82,000 21,500 23,000 22,000 yes yes no yes What type of protein separation techniques might she use to protein X from the other proteins. Give a flow...
SIRJE GEL OBJECTIVE: Partial purification of a protein, isocitrate dehydrogenase (IDH), using DEAE ion exchange chromatography. IDH will be eluted from the column using increasing concentrations of KCI. IDH will be tracked using the IDH activity assay and the extent of purification will be analyzed using SDS PAGE Last week we assayed for the activity of the enzyme, isocitrate dehydrogenase, from E.coli strains carrying a plasmid that over-expresses the E. coli IDH gene. To purify proteins from cel extracts, biochemists...
In order to separate protein X using ion exchange chromatography, I am using a resin with carboxylates attached (pka is 6.0). If I wanted to separate protein X, whose plis 8.0, what is the pH range of the buffer that can be used for ion exchange chromatography with this resin? any pH between 6.0 and 8.0 any pH above 6.0. any pH below 8.0 there is no pH range for this resin and protein X. Which of the following is...
a-d plz
7. Protein purification. As a graduate student, the first protein I purified was RNA polymerase (RNAP) from E. coli- Some physical and chemical properties of Ecoli RNAP Molecular max470,000 g/mol polynentide compasitian (subunits): (50 kDa), k andok a pl = 5.34 substrates: NTPs cofactor: Mg? Purification protocol. E coli cells were broken usine Isozyme, yielding a cellualar extract containing a protcome solution 4M (NHOSO, was added to the cellular extract. A white protein precipitate was formed incuding RNAM...
7. Protein purification. As a graduate student, the first protein I purified was RNA polymerase (RNAP) from E. coli. Some physical and chemical properties of E. coli RNAP Molecular mass = 470,000 g/mol polypeptide composition (subunits): a (50 kDa), B (150 kDa) and 6 (70 kDa) pl = 5.34 substrates: NTPS cofactor: Mg Purification protocol. E. coli cells were broken using lysozyme, yielding a cellualar extract containing a proteome solution. 4M (NH4)2SO4 was added to the cellular extract. A white...
1. A biochemist is attempting to separate a DNA-binding protein (protein X) from other proteins in a solution. Only three other proteins (A, B, and C) are present. The proteins have the following properties: pl (isoelectric point) Size Mr Bind to DNA? protein A 7.4 protein B 3.8 protein C 7.9 protein X7.8 82,000 21,500 23,000 22,000 yes yes no yes What type of protein separation techniques might she use to protein X from the other proteins. Give a flow...
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