Quantitative analysis of protein far UV circular dichroism spectra by neural networks

Böhm, Gerald; Muhr, Rudolf; Jaenicke, Rainer

doi:10.1093/protein/5.3.191

Cited by 1,066 publications

(749 citation statements)

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“…Deconvolution of the nMOMP CD spectra with the CDNN secondary structure prediction algorithm40 predicted the fraction of parallel and anti‐parallel β‐sheet secondary structure elements in C. trachomatis Serovars D, E, F, and J nMOMP to be in average 23.6% ± 0.7%. The fraction of β‐sheet elements of C. muridarum nMOMP was with 36.9% significantly higher.…”

Section: Resultsmentioning

confidence: 99%

“…The CD spectrum we obtained for C. muridarum MOMP had a minimum at 215 nm and a shape that was consistent with the spectrum reported by Sun et al37 We used the CDNN algorithm to deconvolute the CD spectra and estimate the secondary structure composition of all nMOMP preparations (Table 2). 40 The major component of the C. muridarum nMOMP secondary structure was random coil (34.1%), and α‐helix contributed 11.6%. CDNN allows distinguishing between parallel and anti‐parallel β‐sheet secondary structure.…”

Section: Discussionmentioning

confidence: 98%

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Spectroscopic analysis of chlamydial major outer membrane protein in support of structure elucidation

et al. 2018

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Chlamydial major outer membrane protein (MOMP) is the major protein constituent of the bacterial pathogen Chlamydia trachomatis. Chlamydia trachomatis Serovars D–K are the leading cause of genital tract infections which can lead to infertility or ectopic pregnancies. A vaccine against Chlamydia is highly desirable but currently not available. MOMP accounts for ~ 60% of the chlamydial protein mass and is considered to be one of the lead vaccine candidates against C. trachomatis. We report on the spectroscopic analysis of C. trachomatis native MOMP Serovars D, E, F, and J as well as C. muridarum MOMP by size exclusion chromatography multi angle light scattering (SEC MALS), circular dichroism (CD) and attenuated total reflectance Fourier transform infrared spectroscopy (ATR‐FTIR). MOMP was purified from the native bacterium grown in either adherent HeLa cells or in different suspension cell lines. Our results confirm that MOMP forms homo‐trimers in detergent micelles. The secondary structure composition of C. trachomatis MOMP was conserved across serovars, but different from composition of C. muridarum MOMP with a 13% (CD) to 18% (ATR‐FTIR) reduction in β‐sheet conformation for C. trachomatis MOMP. When Serovar E MOMP was isolated from suspension cell lines the α‐helix content increased by 7% (CD) to 13% (ATIR‐FTIR). Maintenance of a native‐like tertiary and quaternary structure in subunit vaccines is important for the generation of protective antibodies. This biophysical characterization of MOMP presented here serves, in the absence of functional assays, as a method for monitoring the structural integrity of MOMP.

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Section: Resultsmentioning

confidence: 99%

Section: Discussionmentioning

confidence: 98%

Spectroscopic analysis of chlamydial major outer membrane protein in support of structure elucidation

et al. 2018

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“…The data were corrected for the baseline con-tribution of the buffer and the observed ellipticities were converted into the mean residue ellipticities (θ) based on a mean molecular mass per residue of 110 Da. Secondary structure was estimated from fi tted far-UV CD spectra using the CDNN software package [38].…”

Section: Circular Dichroism Spectroscopymentioning

confidence: 99%

A 70-kDa molecular chaperone, DnaK, from the industrial bacterium Bacillus licheniformis: gene cloning, purification and molecular characterization of the recombinant protein

et al. 2009

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The heat shock protein 70 (Hsp70/DnaK) gene of Bacillus licheniformis is 1,839 bp in length encoding a polypeptide of 612 amino acid residues. The deduced amino acid sequence of the gene shares high sequence identity with other Hsp70/DnaK proteins. The characteristic domains typical for Hsps/DnaKs are also well conserved in B. licheniformis DnaK (BlDnaK). BlDnaK was overexpressed in Escherichia coli using pQE expression system and the recombinant protein was purifi ed to homogeneity by nickel-chelate chromatography. The optimal temperature for ATPase activity of the purifi ed BlDnaK was 40°C in the presence of 100 mM KCl. The purifi ed BlDnaK had a V max of 32.5 nmol Pi/min and a K M of 439 μM. In vivo, the dnaK gene allowed an E. coli dnaK756-ts mutant to grow at 44°C, suggesting that BlDnaK should be functional for survival of host cells under environmental changes especially higher temperature. We also described the use of circular dichroism to characterize the conformation change induced by ATP binding. Binding of ATP was not accompanied by a net change in secondary structure, but ATP together with Mg 2+ and K + ions had a greater enhancement in the stability of BlDnaK at stress temperatures. Simultaneous addition of DnaJ, GrpE, and NR-peptide (NRLLLTG) synergistically stimulates the ATPase activity of BlDnaK by 11.7-fold.

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“…Background spectra of samples (buffer for protein solutions or liposomes only at concentrations corresponding to those in proteoliposome samples) used for baseline correction. The secondary structure content was calculated by means of JASCO software, by using the program CDNN 2.1 [16].…”

Section: Circular Dichroismmentioning

confidence: 99%

Prion protein structure is affected by pH‐dependent interaction with membranes: A study in a model system

Sesana¹,

Barbiroli²,

Bonomi³

et al. 2007

FEBS Letters

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Interaction of full length recombinant hamster prion protein with liposomes mimicking lipid rafts or non-raft membrane regions was studied by circular dichroism, chemical cross-linking and sucrose gradient ultracentrifugation. At pH 7.0, the protein bound palmitoyloleoylphosphatidylcholine/ cholesterol/sphingomyelin/monosialoganglioside GM1 (GM1) ganglioside liposomes but not palmitoyloleoylphosphatidylcholine alone (bound/free = 0.33 and 0.01, respectively), maintaining the native a-helical structure and monomeric form. At pH 5.0, though still binding to quaternary mixtures, in particular GM1, the protein bound also to palmitoyloleoylphosphatidylcholine (bound/free 0.35) becoming unfolded and oligomeric. The pH-dependent interaction with raft or non-raft membranes might have implication in vivo, by stabilizing or destabilizing the protein.

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Quantitative analysis of protein far UV circular dichroism spectra by neural networks

Cited by 1,066 publications

References 0 publications

Spectroscopic analysis of chlamydial major outer membrane protein in support of structure elucidation

Spectroscopic analysis of chlamydial major outer membrane protein in support of structure elucidation

A 70-kDa molecular chaperone, DnaK, from the industrial bacterium Bacillus licheniformis: gene cloning, purification and molecular characterization of the recombinant protein

Prion protein structure is affected by pH‐dependent interaction with membranes: A study in a model system

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