An Ultracentrifuge Study of the Polymerization of α-Chymotrypsin

Rao, M. S. Narasinga; Kegeles, Gerson

doi:10.1021/ja01554a038

Cited by 147 publications

(93 citation statements)

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“…There is, however, a considerable difference between the cmr shifts of leucylglycine and glycyleucine. Similar behavior has been observed for the proton spectra of peptides containing glycine by Nakamura and Jardetzky.13 It has been reported earlier 14 that the 13 C spectrum of zwitterionic diglycine consists of four peaks, the positions of which differ considerably from those of free glycine. Starting with the chemical shifts of (13) free glycine, we might proceed in the following way to predict the shifts for the glycine carbons in an N-aminoacylglycine (1).…”

Section: Discussionsupporting

confidence: 81%

“…24 The latter possibility has been used to explain the nonequivalence of glycine methylene protons in aminoacylglycines. 13 .The nonequivalence of isopropyl methyl carbons m carbon-13 nmr has been investigated for some alcohols and hydrocarbons 24 ancl the chemical shift difference was found to depend strongly on the steric requirement of the substituents an the carbon to which the isopropyl group is attached, as would indicate differences in conformational populations. In a ~ydrocarbon with one methylene group between the 1s~pro~yl group and the chiral carbon, the methyl sh1ft ddference was 1.0 ppm.…”

Section: S70mentioning

confidence: 99%

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Nuclear magnetic resonance spectroscopy. Carbon-13 chemical shifts of small peptides as a function of pH

Christl¹,

Roberts²

1972

J. Am. Chem. Soc.

144

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Carbon-13 spectra of 27 peptides have been obtained by high-resolution nmr spectroscopy with the aid of proton decoupling. The chemical shifts of the different amino acid moieties are found to have systematic deviations from those of the free zwitterionic amino acids. These effects depend very largely on the position of an amino acid in a peptide, i.e., on whether it is a C-terminal, an N-terminal, or a nonterminal unit, but not on the nature of its neighbors in the chain, unless one of these is proline. The results have substantial value for the determination of amino acid sequences in di-and tripeptides. Prolonation and deprotonation of zwitterionic peptides have considerable effects on the chemical shifts which are, for the most part, limited to the amino acid units directly involved in the reaction. The one important exception to this rule that we have observed is interpreted as being the consequence of a conformational change which the peptides undergo when transformed from the zwitterion to the anion or cation.N umerous publications in recent years emphasize the potential of nmr investigations on amino acids, peptides, and proteins. 3 The application of proton nmr to proteins, however, was shown to be limited because of the large number of different types of protons in a protein combined with a relatively small spread in chemical shifts and large intrinsic line widths. Specific deuteration of proteins results in much simpler proton spectra and allows more detailed studies. The carbon-13 (cmr) spectra of amino acids 4 show much larger chemical shift differences than for the corresponding proton spectra. This fact provides promise with regard to the resolution of specific resonances in the spectra of peptides and proteins. Several recent papers offer results in accord with this prediction. 5 -9 Proteins with specifi.cally labeled carbon-13 amino acids can now be confidently expected to provide detailed information concerning the three-dimensional structure of proteins in solution and interactions of protein with small molecules.We report here the results of a systematic cmr investigation carried out on small peptides with 1 3 C of natural abundance. The objective of this study was to see if cmr spectroscopy could be developed to be useful for the determination of the amino acid sequences in di-and tripeptides, particularly through possible pH dependences of the carbon chemical shifts. , Fellow, 1970, Fellow, -1971 (3) For a review, see G. C. K. Roberts and 0. Jardetzky, Adcan. Protein Chem., 24, 447 (1970 227, 840 (1970).(6) A. Allerhand, D. Doddrell, V. Glushko, D. W. Cochran, E. Wenkert, P. J. Lawson, and F. R. N. Gurd, J. Amer. Chem. Soc., 93, 544 (1971). (7) A. Allerhand, D. W. Cochran, and D. Doddrell, Proc. Nat. Acad.Sei. U. S., 67, 1093 (1970).(8) G. Jung, E. Breitmaier, and W. Voelter, Angew. Chem., lnt. Ed. Eng/., 9, 894 (1970). (9) 42, 1148 (1971). Experimental SectionThe peptides examined in the present research were all commercial materials and were used without further purificat...

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Section: Discussionsupporting

confidence: 81%

Section: S70mentioning

confidence: 99%

Nuclear magnetic resonance spectroscopy. Carbon-13 chemical shifts of small peptides as a function of pH

Christl¹,

Roberts²

1972

J. Am. Chem. Soc.

144

View full text Add to dashboard Cite

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“…It is also reported that above pH = 6, α-chymotrypsin undergoes aggregation of a higher order than the monomer-dimer. (15) In the presence of calcium chloride, all the thermal transitions were irreversible; hence we have reported only the T m values for these cases. Figure 4 shows the alternation of the transition temperature which is widely considered as a protein stability index…”

Section: Surface Tension Measurementsmentioning

confidence: 99%

Thermodynamics of the interactions of calcium chloride with α-chymotrypsin

Kar

Alex

Kishore

2002

The Journal of Chemical Thermodynamics

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“…The principles outlined are being applied to several reversibly polymerizing proteins, among them alpha-chymotrypsin and lobster hemocyanin, for which considerable information is already available (14)(15)(16)(17)(18).…”

Section: Related Workmentioning

confidence: 99%

Z -Average Sedimentation Coefficient in Reversibly Self-Aggregating Systems

Kegeles¹

1972

Proc. Natl. Acad. Sci. U.S.A.

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A theoretical treatment of the determination of the Z-average sedimentation coefficient in reversibly self-aggregating systems is presented. These equations are useful for determination of formation constants of polymers.When "concentration-boundaries" are produced in an ultracentrifuge by layering techniques within synthetic boundary cells (1, 2), gravitationally stable boundaries result in the case of rapidly reversible self-aggregating systems (3,4,16). For such systems, the "concentration-boundary" moves at a rate faster than that of a conventional free boundary below buffer (4). The present analysis shows that in this particular case, the rate of movement of the "concentration-boundary" defines the Z-average sedimentation coefficient, as the concentration difference across the boundary becomes very small. In the case of a monomer in equilibrium with a single higher polymer, the combination of the Z-average and weightaverage sedimentation coefficients leads to a particularly simple method for the evaluation of the formation constant of the polymer. Previous studiesIn their examination of such "concentration-boundaries" in an ultracentrifuge, Schachman and colleagues (3, 4) concentrated primarily on the use of such boundaries as a means for studying hydrodynamic interaction between sedimenting solute species. They, as well as Steiner (16), showed that stable boundaries with sedimentation rates higher than the weight-average sedimentation coefficient occur for rapidly reversible self-aggregating systems, and that such results are consistent with the mass-conservation requirements for solute. Another examination of this problem (5) suggested that a series of separated schlieren peaks could be obtained from such "concentration-boundaries," providing individual data for each species. This result has not been realized in practice. Related workThe study of such "concentration-boundaries" in electrochemistry has provided valuable data on transference numbers (6). In the field of gel-permeation chromatography, Gilbert et al. (7,8) have made excellent progress in their characterization of chemically reacting macromolecular systems. Part of the formulation developed here is similar to that reported (7,8). However, because of the classical use of the ultracentrifuge to characterize parameters of molecular weight distribution functions (9), it is natural, in addition, to examine the type of average sedimentation coefficient ac-2577 [3]The weight average sedimentation coefficient is also defined by (4)The relationship derived from the conservation of mass for the limiting sedimentation coefficient of the "concentrationboundary" as the concentration increment across it approaches zero is given (3, 4) byIn a reversibly self-associating reacting system containing many species, this relation is generalized to [6] SD= d(g.sc)/dc = d(lsici)/dc. i In order to take into account the hydrodynamic concentration dependence of the sedimentation coefficients, it will be assumed that for each species Si = si'/(1 + kc). [7] Here,...

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An Ultracentrifuge Study of the Polymerization of α-Chymotrypsin

Cited by 147 publications

References 2 publications

Nuclear magnetic resonance spectroscopy. Carbon-13 chemical shifts of small peptides as a function of pH

Nuclear magnetic resonance spectroscopy. Carbon-13 chemical shifts of small peptides as a function of pH

Thermodynamics of the interactions of calcium chloride with α-chymotrypsin

Z -Average Sedimentation Coefficient in Reversibly Self-Aggregating Systems

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