Derivation and use of a formula to calculate the net charge of acid-base compounds. Its application to amino acids, proteins and nucleotides

Cameselle, José Carlos; Ribeiro, João Meireles; Sillero, Antonio

doi:10.1016/0307-4412(86)90176-7

Cited by 45 publications

(41 citation statements)

References 5 publications

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“…The molecular mass of the enzyme obtained by both procedures and after elution from the cromatofocusing PBE 94 column was about 52 kDa, indicating that during this step, the complex enzyme-inhibitor dissociated, hence, the similarity of the pI values indicated above. Recently, we have developed methods (8,31) to calculate theoretical pI values of proteins, provided that their amino acid compositions are known. The experimental pl value (about 5.0) differed from the theoretical one (9.7) deduced from the amino acid composition described for the 5'-nucleotidase (24) and calculated as described previously (8,31).…”

Section: Discussionmentioning

confidence: 99%

“…Recently, we have developed methods (8,31) to calculate theoretical pI values of proteins, provided that their amino acid compositions are known. The experimental pl value (about 5.0) differed from the theoretical one (9.7) deduced from the amino acid composition described for the 5'-nucleotidase (24) and calculated as described previously (8,31). This discrepancy can be interpreted as either an indication of the presence of impurities in the enzyme preparation, from which the amino acid composition was determined, or as being due to a particular folding of the protein that hinders part of the positively charged amino acids of the protein.…”

Section: Discussionmentioning

confidence: 99%

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Hydrolysis of bis(5'-nucleosidyl) polyphosphates by Escherichia coli 5'-nucleotidase

et al. 1989

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Two enzymatic activities that split diadenosine triphosphate have been reported in Escherichia coli: a specific Mg-dependent bis(5'-adenosyl) triphosphatase (EC 3.6.1.29) and the bis(5'-adenosyl) tetraphosphatase (EC 3.6.1.41). In addition to the activities of these two enzymes, a different enzyme activity that hydrolyzes dinucleoside polyphosphates is described. After purification and study of its molecular and kinetic properties, we concluded that it corresponded to the 5'-nucleotidase (EC 3.1.3.5) that has been described in E. coli. The enzyme was purified from sonic extracts and osmotic shock fluid. From sonic extracts, two isoforms were isolated by chromatography on ion-exchange Mono Q columns; they had a molecular mass of about 100 kilodaltons (kDa). From the osmotic shock fluid, a unique form of 52 kDa was recovered. Mild heating transformed the 100-kDa isoform to a 52-kDa form, with an increase in activity of about threefold. The existence of a 5'-nucleotidase inhibitor described previously, which associates with the enzyme and is not liberated in the osmotic shock fluid, may have been responsible for these results. The kinetic properties and substrate specificities of both forms (52 and 100 kDa) were almost identical. The enzyme, which is known to hydrolyze AMP and uridine-(5')-diphospho-(1)-alpha-D-glucose, but not adenosine-(5')-diphospho-(1)-alpha-D-glucose, was also able to split adenosine-(5')-diphospho-(5)-beta-D-ribose, ribose-5-phosphate, and dinucleoside polyphosphates [diadenosine 5',5'''-P1,P2-diphosphate,diadenosine 5',5'''-P1,P3-triphosphate, diadenosine 5',5'''-P1,P4-tetraphosphate, and bis(5'-guanosyl) triphosphate]. The effects of divalent cations and pH on the rate of the reaction with different substrates were studied.

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Section: Discussionmentioning

confidence: 99%

Section: Discussionmentioning

confidence: 99%

Hydrolysis of bis(5'-nucleosidyl) polyphosphates by Escherichia coli 5'-nucleotidase

et al. 1989

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“…As expected, the insertion of Asp lowered the pI while Lys increased it. An approximate net charge of the proteins at different pH values can be calculated (Cameselle et al,1996) from the amino acid-residue content of the proteins (Tables I, II). The influence of the ionizable amino acid residues on neighboring residues is not taken into account in such a calculation.…”

Section: Purification and Characterization Of The Z Derivativesmentioning

confidence: 99%

Genetic engineering of protein-peptide fusions for control of protein partitioning in thermoseparating aqueous two-phase systems

Berggren¹,

Veide²,

Nygren³

et al. 1999

Biotechnol. Bioeng.

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Genetic engineering has been used for the fusion of peptides, with different length and composition, on a protein to study the effect on partitioning in aqueous two‐phase systems containing thermoseparating polymers. Peptides containing 2–6 tryptophan residues or tryptophan plus 1–3 lysine or aspartate residues, were fused near the C‐terminus of the recombinant protein ZZT0, where Z is a synthetic IgG‐binding domain derived from domain B in staphylococcal protein A. The partitioning behavior of the peptides and fusion proteins were studied in an aqueous two‐phase system composed of dextran and the thermoseparating ethylene oxide–propylene oxide random copolymer, EO30PO70. The zwitterionic compound β‐alanine was used to reduce the charge‐dependent salt effects on partitioning, and to evaluate the contribution to the partition coefficient from the amino acid residues, Trp, Lys, and Asp, respectively. Trp was found to direct the fusion proteins to the EO–PO copolymer phase, while Asp and Lys directed them to the dextran phase. The effect of sodium perchlorate and triethylammonium phosphate on the partitioning of the fusion proteins was also studied. Salt effects were directly proportional to the net charge of the fusion proteins. Sodium perchlorate was found to be 3.5 times more effective in directing positively charged proteins to the EO–PO copolymer phase compared to the effect of triethyl ammonium phosphate on negatively charged proteins. An empirical correlation has been tested where the fusion protein partitioning is a result of independent contributions from unmodified protein, fused peptide, and salt effects. A good agreement with experimental data was obtained which indicates the possibility, by independent measurements of partitioning of target protein and fusion peptide, to approximately predict the fusion protein partitioning. © 1999 John Wiley & Sons, Inc. Biotechnol Bioeng 62: 135–144, 1999.

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“…However, we must consider the following: the percentage of the net charge dissociated does not remain permanently decoupled in that percentage, but there is a time of molecular fractionation for both the dissociated (t d ) and undissociated (t nd ) form which is equivalent to the degree of molecular dissociated (a) and undissociated (1-a). The expressions once incorporated into the Henderson-Hasselbalch equation and making the corresponding re-arrangement allows to obtain the expressions [3] to calculate the net charge (Z net ) of an acid-base group or a molecule with an infinite number of acid-base groups.…”

Section: Introductionmentioning

confidence: 99%

“…Research have addressed the theoretical calculations of the pI and the net electric charge of proteins and other macromolecules [2][3][4][5][6][7] whose topics have been used to explain to students in regular courses of Biochemistry [8].…”

Section: Introductionmentioning

confidence: 99%

Design of a software for calculating isoelectric point of a polypeptide according to their net charge using the graphical programming language LabVIEW

Tovar

2017

Biochem Molecular Bio Educ

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A software to calculate the net charge and to predict the isoelectric point (pI) of a polypeptide is developed in this work using the graphical programming language LabVIEW. Through this instrument the net charges of the ionizable residues of the polypeptide chains of the proteins are calculated at different pH values, tabulated, pI is predicted and an Excel (-xls) type file is generated. In this work, the experimental values of the pIs (pI) of different proteins are compared with the values of the pIs (pI) calculated graphically, achieving a correlation coefficient (R) of 0.934746 which represents a good reliability for a p < 0.01. In this way the generated program can constitute an instrument applicable in the laboratory, facilitating the calculation to graduate students and junior researchers.

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Derivation and use of a formula to calculate the net charge of acid-base compounds. Its application to amino acids, proteins and nucleotides

Cited by 45 publications

References 5 publications

Hydrolysis of bis(5'-nucleosidyl) polyphosphates by Escherichia coli 5'-nucleotidase

Hydrolysis of bis(5'-nucleosidyl) polyphosphates by Escherichia coli 5'-nucleotidase

Genetic engineering of protein-peptide fusions for control of protein partitioning in thermoseparating aqueous two-phase systems

Design of a software for calculating isoelectric point of a polypeptide according to their net charge using the graphical programming language LabVIEW

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