Calculation and use of protein‐derived conformation‐related spectra for the estimate of the secondary structure of proteins from their infrared spectra

Eckert, K; Grosse, Richard; Malur, J.; Repke, K.

doi:10.1002/bip.1977.360161116

Cited by 41 publications

(36 citation statements)

References 15 publications

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“…10). In this technique, position and shape of the amide I vibration near 1650 cm-1 are monitored in a solution of the protein in 2H20 (70,71). For comparison, spectra of deuterated reference proteins are also included.…”

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confidence: 99%

Structure of Mammalian Metallothionein

Kägi¹,

Vašák²,

Lerch³

et al. 1984

Environmental Health Perspectives

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All mammalian metallothioneins characterized contain a single polypeptide chain of 61 amino acid residues, among them 20 cysteines providing the ligands for seven metalbinding sites. Native metallothioneins are usually heterogeneous in metal composition, with Zn, Cd, and Cu occurring in varying proportions. However, forms containing only a single metal species, i.e., Zn, Cd, Ni, Co, Hg, Pb, Bi, have now been prepared by in vitro reconstitution from the metal-free apoprotein. By spectroscopic analysis of such derivatives it was established that all cysteine residues participate in metal binding, that each metal ion is bound to four thiolate ligands, and that the symmetry of each complex is close to that of a tetrahedron.To satisfy the requirements of the overall Me7(Cys-)20 stoichiometry, the complexes must be combined to form metal-thiolate cluster structures. Experimental proof for the occurrence of such clusters comes from the demonstration of metal-metal interactions by spectroscopic and magnetic means. Thus, in Co(II)7-metallothionein, the Co(II)-specific ESR signals are effectively suppressed by antiferromagnetic coupling of juxtaposed paramagnetic metal ions. By monitoring changes in ESR signal size occurring on stepwise incorporation of Co(II) into the protein, it is possible to follow the building up of the clusters. This process is biphasic. Up to binding of four equivalents of Co(II), the ESR amplitude increases in proportion to the metal content, indicating generation of magnetically noninteracting high-spin complexes. However, upon addition of the remaining three equivalents of Co(II), these features are progressively suppressed, signaling the formation of clusters. The same mode of cluster formation has also been documented for Cd and Hg.The actual spatial organization of the clusters and the polypeptide chain remains to be established. An attractive possibility is the arrangement of the tetrahedral metal-thiolates in adamantane-like structures surrounded by properly folded segments of the chain providing the ligands. 1H-NMR data and infrared absorption measurements are consistent with a tightly folded structure rich in 3-type conformation.

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mentioning

confidence: 99%

Structure of Mammalian Metallothionein

Kägi¹,

Vašák²,

Lerch³

et al. 1984

Environmental Health Perspectives

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“…A maximum at this wavenumber is characteristic for proteins rich in a-helical structure and poor in (or without) /? structure, like myoglobin (171, methemoglobin and cytochrome c [18]. We find a molar absorption per peptide bond at 1650 cm-' of 412 I .…”

Section: Infrared Spectroscopymentioning

confidence: 76%

“…the wavenumber region considered to be characteristic for / I structure in proteins [18,20]. At present we cannot decide whether the minor bands found at 1673 cm-' and 1652 cm-' do have real structural significance.…”

Section: Infrared Spectroscopymentioning

confidence: 99%

“…The major amide I' band of synthetic polypeptides in antiparallel-chain pleated sheets is in general found at clearly smaller wavenumbers [24] than that of proteins very rich in structure or the protein-derived /? reference spectrum [17,18,20]. Sternberg and Thornton [29] find an average number of 4.7 strands per shect upon an analysis of 57 /I-sheet structures from known protein structures.…”

Section: Infrared Spectroscopymentioning

confidence: 99%

“…Assuming that synthetic polypeptides do form very extended fi-sheets, we observe that thc wavenumbers found for the maximum of the amide I' band of gramicidin S (1647 cm-I), the /? reference spectrum derived from several known proteins (1634 cm-') [18] and of the antiparallel structure of poly(L-lysine) (1612 cm-') [22], decrease with increasing number of strands in the corresponding sheets.…”

Section: Infrared Spectroscopymentioning

confidence: 99%

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Secondary Structure of the lac Repressor Headpiece

Schnarr

Maurizot

1982

European Journal of Biochemistry

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The secondary structure of the short tryptic headpiece of the lac repressor has been investigated by the analysis of its infrared and circular dichroic spectra. For the latter we used the method of Provencher and Glockner [Biochemistry (1981) 20, 33-37], which seems to be at present the most successful for the determination of the p content of proteins. Nevertheless our results indicate that in the case of the lac repressor headpiece this method overestimates the amount of [j structure. We find that the headpiece contains an important helical content of about 50:,;, depending slightly on the ionic strength. A decomposition of the infrared spectrum in a sum of Gaussian curves reveals clearly the absence of a vibrational band around 1630 cm-', excluding thus the presence of a multi-stranded 8-pleated sheet. The only structure compatible with the infrared results seems to be a twostranded antiparallel fl sheet, as judged from our results on the fl-sheet model-compound gramicidin s. The unusually strong intensity of the amide I' band is in favour of the existence of such a structure. The quantitative analysis of both infrared and circular dichroism spectra indicates the presence of a certain (but different) amount of fl structure. Comparing these results with several secondary structure predictions, part of the helical residues should be located between Leu-45 and (at least) Arg-35, and an eventual two-stranded j? sheet should be situated in the N-terminal part of the headpiece.In the Escherichia coli lac operon the expression of the structural genes is negatively controlled by the interaction of a protein, the lac repressor, with the luc operator [l, 21. Intact repressor has a molecular weight of 154000 and is a tetramer of four identical subunits of 360 amino acids. Upon limited proteolysis of lac repressor in high-ionic-strength buffer one obtains four small N-terminal 'headpieces' (residues 1 -51, 1 -56 or I -59 depending on the protease used and the duration of hydrolysis) and a tetrameric 'core' [3,4]. There is little doubt now that the headpieces constitute the major, if not the entire, part of the DNA-binding domain of the lac repressor. It has been shown that the isolated headpiece has nonspecific affinity for DNA [3,5,6] and induces the same conformational change as does the intact lac repressor upon nonspecific binding (our unpublished results). Ogata and Gilbert [7,8] have shown that the pattern of modification of the Iuc operator by dimethyl sulfate is very similar whether it is induced by the intact protein or by the headpiece. It is thus of considerable interest to determine the structure of the headpiece in order to understand how it interacts with the DNA.Information on the global shape of the headpiece has recently been obtained by neutron-scattering measurements, suggesting an elongated form with a long dimension of about 5 nm and a cross-sectional diameter of 1.5-2 nm [9]. On the other hand, local information, concerning mainly the four tyrosine residues, has been obtained by 'H and I9F NMR [lo-1...

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Bovine serum albumin observed by infrared spectrometry. I. Methodology, structural investigation, and water uptake

2000

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The results of preliminary infrared (IR) spectrometry experiments on bovine serum albumin (BSA) films are presented. An analysis of spectral variations due to raising the temperature and deuteration of NH groups leads to the assignment of most IR bands of BSA. From this analysis we furthermore deduce that at 115°C only hydrogen bonds established by NH groups on the still present H2O molecules, which are so strongly bound to the protein that they do not evaporate, are weakened, some of which are broken. These NH⋮OH2 groups represent some 5% of all NH groups in the dried protein. Spectral changes due to hydration by water vapor are also analyzed and a precise method to measure the water‐vapor pressure of the atmosphere surrounding the BSA film, or equivalently the relative humidity, is described. Various procedures to measure the number of H2O molecules embedded in BSA are then presented and evaluated. One of them is selected as the best one for proteins, because it matches previous measurements based on gravimetric methods. This procedure is subsequently used in a study that is devoted to the determination of the various hydrogen‐bond configurations, or interaction configurations, which are adopted by H2O molecules during the various steps of hydration of BSA. This first analysis of hydration spectra allows the completion of the assignment of IR bands. The various spectral components of the amide I band, which are interchanged during the hydration process, cannot be assigned to various secondary structures, as is usually proposed. It suggests that this usual assignment should be used with care, especially by taking into account the state of hydration, when one wishes to obtain structural information from it. © 2000 John Wiley & Sons, Inc. Biopolymers (Biospectroscopy) 62: 40–53, 2001

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Calculation and use of protein‐derived conformation‐related spectra for the estimate of the secondary structure of proteins from their infrared spectra

Cited by 41 publications

References 15 publications

Structure of Mammalian Metallothionein

Structure of Mammalian Metallothionein

Secondary Structure of the lac Repressor Headpiece

Bovine serum albumin observed by infrared spectrometry. I. Methodology, structural investigation, and water uptake

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