Ultraviolet spectra Of Proteins and Amino Acids

Wetlaufer, Donald B.

doi:10.1016/s0065-3233(08)60056-x

Cited by 977 publications

(475 citation statements)

References 221 publications

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“…It is clear that these techniques do not always lead to t values in good agreement. The same is true for ribonuclease A (Wetlaufer, 1962) and other proteins (Gill & von Hippel, 1989). In this paper, we report t values determined for RNase T1 and six mutants, and for seven other proteins by the dry weight and/or Edelhoch methods.…”

mentioning

confidence: 78%

“…Similar but less extensive tables were compiled by Wetlaufer (1962), Gill and von Hippel (1989), and Mach et al (1992). These tables all contained errors, and there are surely some errors in our table, but we doubt that they will significantly change the average t values we derive from our analysis.…”

Section: Cn Pace Et Almentioning

confidence: 90%

“…Some of the entries were taken from tables in other papers: 1-1 1 are from Table V in Wetlaufer (1962); 18-36 are from Table 3 in Gill and von Hippel (1989); and 37-47 are from Table 1 in Mach et al (1992). In the latter two tables, we averaged the most reliable values when more than one t value was given.…”

Section: Cn Pace Et Almentioning

confidence: 99%

“…Also, any errors we found in these tables were corrected. In Wetlaufer (1962), for example, the Trp, Tyr, and cystine content was correct for only 1 of the 11 entries, and in Gill and von Hippel (1989), all of the cystine contents were wrong.…”

Section: Cn Pace Et Almentioning

confidence: 99%

“…Here, the Trp estimates d o not differ significantly, but the Tyr estimates differ by almost 4%. Finally, in the last row, we give an analysis of 11 proteins used by Wetlaufer (1962) to first test the possibility of estimating E values for proteins using model compound data. We will see in Table 9 that these t estimates for Trp, Tyr, and cystine are considerably better at predicting t values for a protein than the E values used by Wetlaufer (1962).…”

Section: Cn Pace Et Almentioning

confidence: 99%

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How to measure and predict the molar absorption coefficient of a protein

et al. 1995

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The molar absorption coefficient, E, of a protein is usually based on concentrations measured by dry weight, nitrogen, or amino acid analysis. The studies reported here suggest that the Edelhoch method is the best method for measuring E for a protein. (This method is described by Gill and von Hippel [1989, Anal Biochem 182:319-3261 and is based on data from Edelhoch [ , Biochemistry 6:1948.) The absorbance of a protein at 280 nm depends on the content of Trp, Tyr, and cystine (disulfide bonds). The average E values for these chromophores in a sample of 18 well-characterized proteins have been estimated, and the E values in water, propanol, These ~(280) values are quite reliable for proteins containing Trp residues, and less reliable for proteins that do not. However, the Edelhoch method is convenient and accurate, and the best approach is to measure rather than predict E .

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mentioning

confidence: 78%

Section: Cn Pace Et Almentioning

confidence: 90%

Section: Cn Pace Et Almentioning

confidence: 99%

Section: Cn Pace Et Almentioning

confidence: 99%

Section: Cn Pace Et Almentioning

confidence: 99%

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How to measure and predict the molar absorption coefficient of a protein

et al. 1995

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Overview of the Characterization of Recombinant Proteins

Denslow

Wingfield

Rose³

1995

CP Protein Science

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This overview provides guidelines for the characterization of recombinantly expressed proteins (e.g., verifying primary structure and appropriate post-translational modifications), along with methodologies for characterizing the proteins according to size, X-ray structure, absorbance, biological activity, and subunit structure. A flow chart presents a suggested path for fully characterizing recombinant protein and involves equipment for HPLC, mass spectrometry, circular dichroism, NMR and fluorescence spectroscopy. Also covered are sources and consequences of contamination in protein solutions.

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Determining the Identity and Purity of Recombinant Proteins by UV Absorption Spectroscopy

1995

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Because each protein (gene product) has a unique amino acid sequence, the particular aromatic amino acid content of each protein results in a unique spectrum in the near-UV (250 to 350 nm) region. The highly specific microenvironment experienced by each aromatic residue in the three-dimensional protein matrix results in fine shifts in a protein's spectrum. This unit provides protocols for the detection and analysis of UV spectra of recombinant proteins and their peptide fragments. The unique UV spectral properties of proteins can in turn be used to assess their purity. This application is inherent in the use of a diode array detector to monitor the effluent from a high-performance liquid chromatography (HPLC) column. A protocol using this technique to assess the purity of recombinant proteins is presented.

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Ultraviolet spectra Of Proteins and Amino Acids

Cited by 977 publications

References 221 publications

How to measure and predict the molar absorption coefficient of a protein

How to measure and predict the molar absorption coefficient of a protein

Overview of the Characterization of Recombinant Proteins

Determining the Identity and Purity of Recombinant Proteins by UV Absorption Spectroscopy

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