Proteínas de Ferro-enxofre

Postagem para reunir artigos e referências sobre proteínas de ferro-enxofre, além dos casos especiais da cadeia de transporte de elétrons mitocondrial, fotossíntese, etc.

  • Para quem ficou impressionado (eu fiquei!) com o “cabo” de 7 a 9 clusters de [FeS] presente no Complexo I, vejam esta enzima com 46 clusters [FeS]! A estrutura contém outras curiosidades como metais de tungstênio, canais para transporte de gases e hidratados, e grupos prostéticos incomuns! Alguém maluco o suficiente para modelar esta belezinha???
  • Estruturas cristalográficas de resolução ultra-aumentada permitem a “observação” de átomos de hidrogênio, pares eletrônicos e até, pasmen, densidade de carga de orbitais de fronteira. Vejam este artigo descrevendo uma proteína de [4Fe-4S] com resolução de 0.48 angstroms! Alguém interessado em construir um modelo híbrido QC/MM para esta proteína e verificar a precisão dos cálculos na reprodução da densidade de carga?


G Protein-Coupled Receptors

Precision vs Flexibility in GPCR signaling

Nelson Orsalino Neto Schuback, 9010821, CCM-T25

This post consists in a review of  “Precision vs Flexibility in GPCR signaling, Matthias Elgeti et al.”.It is suitable to highlight that this review is focussed on expose the methodology, the results of the paper and develop further discussions to other students.


Communication between different cells of an organism is essential to it’s own existence and operation. Cells must operate differently according to the organism’s condition, the only way it is possible is with cellular signaling. The mecanism behind cellular signaling is the secretion and acceptance of substances from cell to cell, this substances may be classified between three important classes: Hormones, Neurotransmitters and Citokynes. The signal is recognized by receptor proteins (intracellular or membrane proteins), generally this receptors are membrane bounded or integrated, so they can transduce signals from substances that do not cross the cellular membrane. One of the most important classes of membrane protein receptors is the G Protein-Coupled Receptors, GPCRs for the intimate. The vision is due to this receptors, specifically Rhodopsin, a GPCR that can detect signals indicating the presence of photons. As well, they are involved in many diseases, and are also the target of approximately 40% of all modern medicinal drugs, therefore consisting a great pillar of pharmaceutical industry.

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Structure and Mechanism

GPCRs constitute a large family of membrane protein receptors. Coupling with G Proteins, they are called seven-transmembrane receptors because they pass through the cell membrane seven times. The G protein–coupled receptor is activated by an external signal in the form of a ligand or other signal mediator. This creates a conformational change in the receptor, causing activation of a G protein. Further effect depends on the type of G protein. G proteins are subsequently inactivated by GTPase activating proteins, known as RGS proteins. There are two principal signal transduction pathways involving the G protein–coupled receptors: the cAMP signal pathway and the phosphatidylinositol signal pathway.

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Rhodopsin is light sensitive GPCR  involved in visual phototransduction. The cofactor Retinol is produced in the retina from Vitamin A, from dietary beta-carotene. Isomerization of 11-cis-retinal into all-trans-retinal by light induces a conformational change (bleaching) in opsin, continuing with metarhodopsin II, which activates the associated G protein transducin and triggers a Cyclic Guanosine Monophosphate, second messenger, cascade. Metarhodopsin II activates the G protein transducin (Gt) to activate the visual phototransduction pathway. When transducin’s α subunit is bound to GTP, it activates cGMP phosphodiesterase. cGMP phosphodiesterase hydrolyzes cGMP (breaks it down). cGMP can no longer activate cation channels. This leads to the hyperpolarization of photoreceptor cells and a change in the rate of transmitter release by these photoreceptor cells.

Experimental Techniques

Techniques involved in the experiment aim to study the conformational diversity of rhodopsin in membrane environment and extend the static picture provided by the available crystal structures.

- FTIR spectroscopy:

Infrared spectroscopy is a technique in which infrared electromagnetic waves are measured in order to identify compounds, generally organic. Infrared spectroscopy exploits the fact that molecules absorb specific frequencies that are characteristic of theirstructure, these absorptions are resonant frequencies. Fourier transform infrared (FTIR) spectroscopy is a measurement technique that guides the radiation throught an interferometer and applies Fourier transformation to obtain the spectrum.

- Molecular Dynamics (MD):

Molecular dynamics (MD) is a computer simulation method for studying the physical movements of atoms andmolecules, and is thus a type of N-body simulation.  The atoms and molecules are allowed to interact for a fixed period of time, giving a view of the dynamical evolution of the system. Because molecular systems typically consist of a vast number of particles, it is impossible to determine the properties of such complex systems analytically; MD simulation circumvents this problem by using numerical methods.

In the article, FTIR spectroscopy of the rhodopsin in the absence or presence of GαCT and GγCT, this experiment aimed to identify the binding modes of rhodopsin and these peptides. Furthermore, MD simulations were performed using rhodopsin crystal structures in order to study the flexibility of CL3, specially when GαCT and GγCT are interacting with the opsin.

Experimental Results

The experiment section was divided into two parts, first experiments involving FTIR spectroscopy and then experiments involving MD simulations. Each experiment takes a step forward to understand more the structure and the mechanism of Rhodopsin.

- (FTIR spectroscopy) Analysis of metarhodopsin states in equilibrium in a pH range: 

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The first experiment aimed to analyse the equilibrium between metastates of rhodopsin (MI, MIIa, MIIb, IIbH+) (Scheme 1) in a variance of pH. The technical data are listed above in the image caption, after choosing the wavelength to analyse the fraction of metastates  against pH variance. The result was plotted with Henderson-Hasselbalch equation plot for comparison proposes.

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(FTIR spectroscopy) Analysis of G Protein C-terminal interaction with metarhodopsin in a pH range: 

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This second experiment aimed to analyse the equilibrium between metastates of rhodopsin (MI, MIIa, MIIb, IIbH+), as before, but now interacting the rhodopsin with GαCT and GγCT. This experiment shows how G protein binding can stabilize rhodopsin’s structure in several pH. As Figure 2 shows, GαCT stabilizes MIIbH+ at higher pH (~pH9) and GγCT stabilizes MIIb and MIIbH+, but is less effective in the protonated form than GαCT.

(FTIR spectroscopy) Analysis of G Protein C-terminal binding influence in rhodopsin receptor structure: 

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This part of the experiment shows in Figure 3 how GαCT and GγCT binding change receptor structure. In Figure 3A the Protein Binding Spectra (PBS) of GαCT, with it’s high PBS values, is showed to has a high influence on receptor structure. In Figure 3B, PBS of GγCT tells that it has influence, but it is not so strong as GαCT interaction. Finally in Figure 3C, the mutant K231A interaction with GαCT is compared with the normal rhodopsin, the result shows it is not so different. It is important to highlight the shift in molecular vibrations after protonation, that is why the analysis takes data on spectra obtained after H2O/2H2O exchange.

(MD simulations) Analysis of GαCT-CL3 interaction:

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The 200-400 ns simulation of Cl3-GαCT interaction shows in three different situations (A- inactive, B- active, C- active binded to GαCT) how CL3 is flexible. The results A and B shows the high freedom degree of movement of CL3. In the result C, the CL3 is still the most flexible, althought it is much more stable than in the other analysed situations.

PS: It is important to highlight that the experiment arrangement data is all described in the figure captions.

- Discussion in the Free Energy Diagram of MI, MIIa, MIIb and MIIbH+:

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 The schematic free energy landscape illustrating the change of receptor conformations along the thermal activation path under physiological conditions showed that the energy of each metastate of rhodopsin and how binding GγCT and GαCT lowers it’s energy. In Figure 5A, GγCT binds almost equally to MIIb and MIIbH+, but the same does not occur with GαCT, which interaction lowers more the MIIbH+ energy.


The article shows that Rhodopsin’s perfect switching function depends on two factors: signal fidelity and speed. Fidelity relies in the first instance on the inactivating and activating retinal ligands which are covalently bound to the receptor and sufficiently potent to shift the equilibria of receptor conformations to the inactive or active side. Equally important, however, is an accurate and productive coupling mechanism of the activated receptor and G protein.

Blog sobre biofísica estatística

Para interessados em simulação computacional de biomoléculas, o blog Statistical Biophysics vale uma visita. O site é mantido pelo Prof. Daniel Zuckerman que também é autor dos ótimos livros “Statistical Physics of Biomolecules An Introduction” em papel e do online “Physical Lens on the Cell“, que tenta desmistificar alguns dos princípios físicos do funcionamento celular.

Qual o mecanismo de ação de proteínas anti-congelamento?

Diversas proteínas são conhecidas desde os anos 1960 como inibidoras da formação de cristais de gelo em água. Suas estruturas terciárias são diferentes, assim como seus aparentes modos de ligação e até os organismos em que são encontrados. Então, fica a pergunta: qual(is) seu(s) mecanismo(s) microscópico(s) de ação? Acredito que esta fascinante pergunta pode ser atacada por uma combinação de simulação computacional, com amostragem aumentada para variáveis coletivas de nucleação, e teoria de formação de vidros.

Dados experimentais razoavelmente precisos estão disponíveis agora para uma série destas proteínas. Alguém se habilita a estudá-las?