6 research outputs found
Computational Study of the Fe(CN)<sub>2</sub>CO Cofactor and Its Binding to HypC Protein
In
the intricate maturation process of [NiFe]-hydrogenases, the
FeÂ(CN)<sub>2</sub>CO cofactor is first assembled in a HypCD complex
with iron coordinated by cysteines from both proteins and CO is added
after ligation of cyanides. The small accessory protein HypC is known
to play a role in delivering the cofactor needed for assembling the
hydrogenase active site. However, the chemical nature of the FeÂ(CN)<sub>2</sub>CO moiety and the stability of the cofactor–HypC complex
are open questions. In this work, we address geometries, properties,
and the nature of bonding of all chemical species involved in formation
and binding of the cofactor by means of quantum calculations. We also
study the influence of environmental effects and binding to cysteines
on vibrational frequencies of stretching modes of CO and CN used to
detect the presence of FeÂ(CN)<sub>2</sub>CO. Carbon monoxide is found
to be much more sensitive to sulfur binding and the polarity of the
medium than cyanides. The stability of the HypC–cofactor complex
is analyzed by means of molecular dynamics simulation of cofactor-free
and cofactor-bound forms of HypC. The results show that HypC is stable
enough to carry the cofactor, but since its binding cysteine is located
at the N-terminal unstructured tail, it presents large motions in
solution, which suggests the need for a guiding interaction to achieve
delivery of the cofactor
Annexin V staining of PBMCs.
<p>PBMCs were left unstimulated or treated with iron-catechol alone or with PMA alone or in the presence of <i>apo</i>- or <i>holo</i>-Bos d 5. Living cells were gated on forward and side scatter and then plotted for CD4 and Annexin V. (A) Representative pictograms of PBMCs plotted for CD4 and Annexin V. (B) Summary of percentage of CD4+Annexin V+ cells. (C) Summary of percentage of CD4-Annexin V+ cells. Statistical analyses were conducted with repeated measures ANOVA following Newman-Keuls Multiple Comparison test. ***P<0.001, *P<0.05.</p
<i>Apo</i>-, but not <i>holo</i>-Bos d 5 promotes CD4 cells.
<p>PBMCs were treated with PMA alone or in the presence of <i>apo</i>- or <i>holo-</i>Bos d 5. (A) Living cells were gated on forward and side scatter, before gating on CD3 positive cells were performed. CD3 gated cells were then analyzed for their CD4 and CD8 expression.Representative pictograms of CD3 gated PBMCs plotted for CD4 and CD8 (B) Percentage of CD3+CD4+ cells. (C) Percentage of CD3+CD8+ cells. Statistical analyses were conducted with repeated measures ANOVA following Newman-Keuls Multiple Comparison test. **<i>P</i><0.01, ****<i>P</i><0.0001, ns not significant.</p
Bos d 5 is able to bind iron via siderophores.
<p>(A) Bos d 5 (1GX9) with Fe(DHBA)<sub>2</sub> (B) chemical structure of siderophores (C) UV-VIS spectra of Fe(DHBA) complex in the presence or absence of Bos d 5 or with Bos d 5 alone. (D) Iron-staining of <i>apo</i>- or <i>holo</i>-Bos d 5 as well as controls (buffer and iron-siderophore-complex) dotted on nitrocellulose-membrane.</p
Bos d 5 devoid of iron promotes IL13 and IFNγ secretion.
<p>(A) IL13 levels and (B<b>)</b> IFNγ levels of stimulated PBMCs. Statistical analyses were conducted with repeated measures ANOVA following Newman-Keuls Multiple Comparison test. **<i>P</i><0.01, *** P<0.001, ns not significant.</p
Calculated affinities energies and dissociation constants of ligands to Bos d 5.
<p>Calculated affinities energies and dissociation constants of ligands to Bos d 5.</p