title: A unified method for purification of basic proteins.
abstract: Protein purification is still very empirical, and a unified method for purifying proteins without an affinity tag is not available yet. In the postgenomic era, functional genomics, however, strongly demands such a method. In this paper we have formulated a unique method that can be applied for purifying any recombinant basic protein from Escherichia coli. Here, we have found that if the pH of the buffer is merely one pH unit below the isoelectric point (pI) of the recombinant proteins, most of the latter bind to the column. This result supports the Henderson-Hasselbalch principle. Considering that E. coli proteins are mostly acidic, and based on the pI determined theoretically, apparently all recombinant basic proteins (at least pI-1 > or = 6.94) may be purified from E. coli in a single step using a cation-exchanger resin, SP-Sepharose, and a selected buffer pH, depending on the pI of the recombinant protein. Approximately, two-fifths of human proteome, including many if not all nucleic acid-interacting proteins, have a pI of 7.94 or higher; virtually all these 12,000 proteins may be purified using this method in a single step.
authors.affiliation: Department of Oncology, Lombardi Comprehensive Cancer Center, Georgetown University Medical Center, Washington, DC 20057, USA. sa354@georgetown.edu
authors.forename: Sanjay
authors.initials: S
authors.lastname: Adhikari
authors.forename: Praveen Varma
authors.initials: PV
authors.lastname: Manthena
authors.forename: Kamal
authors.initials: K
authors.lastname: Sajwan
authors.forename: Krishna Kiran
authors.initials: KK
authors.lastname: Kota
authors.forename: Rabindra
authors.initials: R
authors.lastname: Roy
mesh_terms: D000818:Animals; D002021:Buffers; D002852:Chromatography, Ion Exchange / Q000379:methods*; D004926:Escherichia coli / Q000378:metabolism; D006801:Humans; D006863:Hydrogen-Ion Concentration; D007526:Isoelectric Point; D051379:Mice; D009696:Nucleic Acids / Q000737:chemistry; D011994:Recombinant Proteins / Q000235:genetics / Q000302:isolation & purification* / Q000378:metabolism
chemical_list: D002021:Buffers; D009696:Nucleic Acids; D011994:Recombinant Proteins