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If a protein with pi of 6 is purified on a DEAE column is it better to use pH 4ph 6 of pH 8 why?
Wiki User
∙ 14y ago
DEAE columns contain a positively charged resin to which negatively charged molecules and proteins will bind. In protein purification, one will generally have the target protein bind the column so the non-target proteins will "flow through" after which the bound proteins are "washed off" the column by changing either the pH or salt concentration. Since the pI of the protein is 6.0, at pH=6.0, the protein has a net charge of zero and will not bind the column, so this pH is not suggested. When the pH is greater than the pI, the protein has a positive charge, so at pH=8.0 the protein will be repulsed by the positively charged resin and will not bind, so pH=8.0 is also not recommended. When the pH is less than the pI, the protein carries a negative charge and will bind the DEAE column and can thus be purified, so the pH=4.0 condition will be productive toward protein purification for this protein.
Wiki User
∙ 14y ago
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How DEAE-Sepharose chromatography causes separation of proteinsDobby and Kreacher?
sum1 help these Kingston university flow of genetics students lol
What does the latin word deam mean?
The word deam means goddess.Latin- deam (Accusative Singular of dea, deae, f)English - "goddess"
What has the author Syed Sajjad Abid Jafri written?
Syed Sajjad Abid Jafri has written: 'Chromatographic behaviour of certain Gram-negative bacteria on DEAE-cellulose'
How do you say 'both the farmer and the sailor heard the reputation of the goddess' in Latin?
Famam deae et agricola et nauta exaudiverunt.The word order is variable without changing the meaning and various other verbs could be used for "they heard": audiverunt, auscultaverunt and others.
What does Macbeth mean by be innocent of the knowledge dearest chucktill thou applaud the deae?
In this passage, Macbeth is telling Lady Macbeth to remain ignorant of his plan to kill King Duncan until after the act is done and she can show approval and praise for it. He wants her to remain innocent of the knowledge of their murderous plot until it is accomplished.
How do you put the spark plug wires back in the correct order on a 1987 Lincoln?
You dont say what engine you have but this is the most commmon. The cylinders are numbered 1-2-3-4 front to back on the pass. side and 5-6-7-8 on the drivers. Pull the #1 plug and bump the starter and when you feel the piston come up on compression, turn the engine slowly intil the timing mark is on top deae center. Pull the cap and where the rotor is pointing is where #1 wire goes. Then contimue with the wires. 1-5-4-2-6-3-7-8 in a counterclockwise direction. #1 wire should be just to the right of center in the back when viewed from the front of the engine unless it has been changed.
Is ion-exchange chromatography is reverse phase chromatography?
The principal of ion exchange chromatography is the most popular method for purifying protein compounds.Other charged molecules are also called an ion exchange if of a protein nature.[Fig 1. Principle of anionic and cationic exchangers.]Biomolecular binding strength depends on solution pH, since it affects the number of ions available for exchange. Proteins can be zwitterions, so you'll need to use either an anionic or cationic exchanger. Whatever the conditions, we have to determine the isoelectric point of the protein at different pHs to see what charges the proteins can have and where it's electroneutral. When pH < pI of a given molecule, it will be positively charged and we'll need a cation exchanger; vice versa when pH > pI. The amphoteric character (ability to react as either acid or base) of proteins allows us to detect ionic interference of other substances and improve our own protein separation. The principle of ion exchange processes will be explained more clearly using an anionic exchange as an example. Today, most people use synthetic organic ion exchangers on a polystyrene base (DOWEX) or natural polymers like cellulose, dextran, or silicate. The macromolecules of the ion exchanger normally make up a 3D network, onto whose surface a huge number of ionizable groups are covalently bonded. Whereas the type of matrix material is generally flow characteristic (the type of ions used and their chemical/mechanical stability are more solid), the groups covalently-bonded to the matrix and the strength of those bonds determine what the exchangable ions can be: every group gives an exchange of very basic (anion exchanger) for very acidic (cation exchanger) character.[Fig 2. Principle of an ion exchange process.]Typical functional groups in an anion exchanger are quaternary amines such as diethyl aminoethyl groups (DEAE - non-denaturing, sorbents have good loading capacity), while those for cation exchangers include organic and inorganic acids like carboxymethyl groups (CMs) or sulfonates. These groups are covalently coupled to the matrix material (Fig. 3). Since exchange groups are only inserted in their ionic form, it's important to know their pK values. Such values can be found by a simple titration curve, as shown for a CM sephadex in Fig 4. Because so many biologically important substances contain ionizable functional groups (amino acids, proteins, nucleotides, nucleic acids, metabolites, etc), biochemical methods for the isolation and separation of charged compounds are quite valuable - some charged compounds are electrostatically bound to the exchanger and others are not.
What are the main types of Fermenters bioreactors used in animal cell culture What are the problems one would face with animal cell culture compared to microbial cell culture?
- Animal cells are large (10 to 20 micrometer diameter), slow growing, and are very shear sensitive. Hence, animal cell reactors must be gently agitated and aerated - Most animal cells are anchorage dependent, hence, surfaces of glass, specially treated plastics, natural polymers such as collagen, or other support materials are used. - Product concentration is very low and toxic metabolites such as ammonium and lactate are produced. Hence suitable measures must be taken to remove these products continuously. In lab scale, T- flasks , spinner bottles, roller bottles and trays containing shallow liquid cultures are used. Large scale reactors: microcarrier systems, hollow fiber reactors, ceramic matrix systems, weighted porous beads) are used for anchorage dependent cells and Stirred tank reactors and bubble column reactors are used for suspension cultures. 1) With roller bottles, the liquid covers about 25% of the surface area. Bottles are rotated about the long axis with the cells adhered to it's sides. They are therefore dipped in the medium for 25 % of the time and are aerated for 75% of the time. It has the advantage of increased surface area for the cells compared to the T-flasks. But it requires high labour and there is bottle to bottle variability. 2) The use of microcarriers of DEAE or dextran for anchorage dependent cells is an attractive approach. They allow high surface to volume ratio. Cells grow on the surface of the microcarriers, usually in the form of monolayers and sometimes as multilayers. But bead to bead contact will expose cells to great stress. Hence microporous microcarriers are used in which cells grow inside them and stress is reduced. 3) For suspension cultures, conventional bioreactors have been modified to reduce shear rates on cells in suspension. Sail-type and axial hydrofoil agitators have been developed and used for suspension cultures. Many shear protection agents lide serum or Pluronic F-68 are added. 4) Hollow fiber reactors are also used to provide a high growth surface- volume ratio and, therefore, high cell concentrations. Cells are immobilized on the external surfaces of hollow fibres, and nutrients pass through the tubes. However the control of microenvironment conditions are different. Other immobilization based reactors: Tubular ceramic matrix reactors , microencapsulation in spherical membranes and gel encapsulation
What is the label for phosphate?
Abbreviations A: Adenosine Ade: Adenine ATP: Adenosine triphosphate B: Nucleobase BMF4TPA: Bis(difluoromethylene)triphosphoric acid BMT: Bismethylene triphosphate Boc: Tert-Butyloxycarbonyl Bop: Bis(2-oxo-3-oxazolidinyl)phosphinic BP : Protected nucleobase BTT: 5-Benzylthio-1-H-tetrazole C: Cytosine CDI: Carbodiimidazole CE: β-Cyanoethyl CEM: Cyanooxymethyl CMPT: N-(cyanomethyl)pyrrolidinium triflate CPG: Controlled pore glass CTP: Cytidine triphosphate Cyt: Cytidine DBU: 1,8-Diazabicyclo[5.4.0]undec-7-ene DCA: Dichloroacetic acid DCI: 4,5-Dicyanoimidazole DEAE: Diethylaminoethyl DIAD: Diisopropyl azodicarboxylate DIPEA: Diisopropylethylamine DMAN: 1,8-Bis-(dimethylamino)naphthalene DMF: N,N-dimethylformamide DMS: Dimethylsulfide DMTr: 4,4′-Dimethoyxltrityl DTD: N,N-dimethylthiuram disulfide EC50 : Half maximal effective concentration EDC/EDCI: 1-Ethyl-3-(3-dimethylaminopropyl)carbodiimide ETT: 5-(Ethylthio)-1H-tetrazole Fm: 9-Fluorenylmethyl Fmoc: Fluorenylmethyloxycarbonyl G: Guanosine Gua: Guanine IC50 : Half maximal inhibitor concentration IEX-HPLC: Ion-exchange high performance liquid chromatography KHMDS: Hexamethyldisilazide LTMPA: Lithium 2,2,6,6-tetramethylpiperidine amide NHS: N-hydroxysuccinimide NMP: Nucleoside monophosphate Npn : Nucleoside polyphosphate NpnN: Dinucleotide polyphosphate NPP: Nucleotide pyrophosphatase/phosphodiesterase Ns: Nosyl NTP: Nucleoside triphosphate NTP: Nucleoside triphosphate Nuc: Nucleotide or nucleoside ODN: Oligodeoxynucleotides ORN: Oligoribonucleotide OTP: Oxathiaphospholane PEP: Phosphoenolpyruvate Pip: Piperidine PK: Pyruvate dinase ppGpp: Guanosin-3′,5′-bispyrophosphate ppp: RNA 5′-triphosphate RNAs PRR: Pattern recognition receptors Py: Pyridine RP18: Reverse phase C18 RSH: RelA-SpoT homolog SAX: Strong anion exchange T: Thymine TBAF: Tetrabutylammonium fluoride TBHP: tert-Butylhydroperoxide TBS: tert-Butyldimethylsilyl TEA: Triethylamine TEAB: Triethylammonium bicarbonate Tf: Trifluoromethylsulfonyl THF: Tetrahydrofuran Thy: Thymidine TMS: Trimethylsilyl Tr: 2,4,6-Triisopropylbenzenesulfonyl Ts: p-Toluenesulfonyl U: Uridine Ura: Uracil UTP: Uridine triphosphate
