Periorbital necrotising fasciitis secondary to scratching. / Fascitis necrosante periorbitaria secundaria a rascado.

An 80 year-old woman with no relevant medical history, consulted for worsening of right palpebral itching and pain after an insect bite. Her eyelids had areas of laceration due to scratching, which rapidly progressed to fibrinoid necrosis over the first 24hours. Lesions were cultivated, revealing Streptococcus pyogenes and Staphylococcus aureus. The patient was admitted to hospital with the diagnosis of periorbital necrotising fasciitis, in order to receive treatment with intravenous ceftriaxone, linezolid, and immediate surgical debridement. She remained in hospital for 17 days. Daily wound management consisted of debridement of necrotic remains, disinfection with chlorhexidine, and wound dressing with mupirocin, sulfadiazine, and miconazole ointments. The patient suffered streptococcal toxic shock syndrome, but she recovered over the first week. Palpebral reconstruction was performed on day 15, consisting of a preauricular total thickness skin graft for the superior eyelid, and lateral malar advancement to cover the lower eyelid. Adequate cosmetic and functional results were obtained.

Tocilizumab as first treatment option in optic neuropathy secondary to Graves' orbitopathy.

Dysthyroid optic neuropathy (DON) is one of the complications that can affect Graves' orbitopathy (GO) patients. Its prevalence is estimated at less than 5%. It is usually treated with intravenous steroids, radiotherapy or orbital decompression. Tocilizumab has been proposed as a treatment option in cases of GO refractory to steroid treatment, with good clinical results. Our aim is to report the case of a patient with optic neuropathy secondary to GO treated with tocilizumab as primary treatment option.

Understanding the molecular basis of the high oxygen affinity variant human hemoglobin Coimbra.

Human hemoglobin (Hb) Coimbra (ßAsp99Glu) is one of the seven ßAsp99 Hb variants described to date. All ßAsp99 substitutions result in increased affinity for O2 and decreased heme-heme cooperativity and their carriers are clinically characterized by erythrocytocis, caused by tissue hypoxia. Since ßAsp99 plays an important role in the allosteric α1ß2 interface and the mutation in Hb Coimbra only represents the insertion of a CH2 group in this interface, the present study of Hb Coimbra is important for a better understanding of the global impact of small modifications in this allosteric interface. We carried out functional, kinetic and dynamic characterization of this hemoglobin, focusing on the interpretation of these results in the context of a growth of the position 99 side chain length in the α1ß2 interface. Oxygen affinity was evaluated by measuring p50 values in distinct pHs (Bohr effect), and the heme-heme cooperativity was analyzed by determining the Hill coefficient (n), in addition to the effect of the allosteric effectors inositol hexaphosphate (IHP) and 2,3-bisphosphoglyceric acid (2,3-BPG). Computer simulations revealed a stabilization of the R state in the Coimbra variant with respect to the wild type, and consistently, the T-to-R quaternary transition was observed on the nanosecond time scale of classical molecular dynamics simulations.

Steered Molecular Dynamics Methods Applied to Enzyme Mechanism and Energetics.

One of the main goals of chemistry is to understand the underlying principles of chemical reactions, in terms of both its reaction mechanism and the thermodynamics that govern it. Using hybrid quantum mechanics/molecular mechanics (QM/MM)-based methods in combination with a biased sampling scheme, it is possible to simulate chemical reactions occurring inside complex environments such as an enzyme, or aqueous solution, and determining the corresponding free energy profile, which provides direct comparison with experimental determined kinetic and equilibrium parameters. Among the most promising biasing schemes is the multiple steered molecular dynamics method, which in combination with Jarzynski's Relationship (JR) allows obtaining the equilibrium free energy profile, from a finite set of nonequilibrium reactive trajectories by exponentially averaging the individual work profiles. However, obtaining statistically converged and accurate profiles is far from easy and may result in increased computational cost if the selected steering speed and number of trajectories are inappropriately chosen. In this small review, using the extensively studied chorismate to prephenate conversion reaction, we first present a systematic study of how key parameters such as pulling speed, number of trajectories, and reaction progress are related to the resulting work distributions and in turn the accuracy of the free energy obtained with JR. Second, and in the context of QM/MM strategies, we introduce the Hybrid Differential Relaxation Algorithm, and show how it allows obtaining more accurate free energy profiles using faster pulling speeds and smaller number of trajectories and thus smaller computational cost.

Binding of the substrate UDP-glucuronic acid induces conformational changes in the xanthan gum glucuronosyltransferase.

GumK is a membrane-associated glucuronosyltransferase of Xanthomonas campestris that is involved in xanthan gum biosynthesis. GumK belongs to the inverting GT-B superfamily and catalyzes the transfer of a glucuronic acid (GlcA) residue from uridine diphosphate (UDP)-GlcA (UDP-GlcA) to a lipid-PP-trisaccharide embedded in the membrane of the bacteria. The structure of GumK was previously described in its apo- and UDP-bound forms, with no significant conformational differences being observed. Here, we study the behavior of GumK toward its donor substrate UDP-GlcA. Turbidity measurements revealed that the interaction of GumK with UDP-GlcA produces aggregation of protein molecules under specific conditions. Moreover, limited proteolysis assays demonstrated protection of enzymatic digestion when UDP-GlcA is present, and this protection is promoted by substrate binding. Circular dichroism spectroscopy also revealed changes in the GumK tertiary structure after UDP-GlcA addition. According to the obtained emission fluorescence results, we suggest the possibility of exposure of hydrophobic residues upon UDP-GlcA binding. We present in silico-built models of GumK complexed with UDP-GlcA as well as its analogs UDP-glucose and UDP-galacturonic acid. Through molecular dynamics simulations, we also show that a relative movement between the domains appears to be specific and to be triggered by UDP-GlcA. The results presented here strongly suggest that GumK undergoes a conformational change upon donor substrate binding, likely bringing the two Rossmann fold domains closer together and triggering a change in the N-terminal domain, with consequent generation of the acceptor substrate binding site.

Molecular mechanism of myoglobin autoxidation: insights from computer simulations.

Myoglobin (Mb) and hemoglobin have the biological ability to carry/store oxygen (O2), a property which requires its heme iron atom to be in the ferrous--Fe(II)--state. However, the thermodynamically stable state in the presence of O2 is Fe(III) and thus the oxidation rate of a globin is a critical parameter related to its function. Mb has been extensively studied and many mutants have been characterized regarding its oxygen mediated oxidation (i.e., autoxidation) rates. Site directed mutants in residues 29 (B10), which shapes the distal cavity, and 64 (E7), the well-known histidine gate, have been shown to display a wide range of autoxidation rate constants. In this work, we have thoroughly studied the mechanism underlying the autoxidation process by means of state-of-the-art computer simulation methodologies, using Mb and site directed mutants as benchmark cases. Our results explain the observed autoxidation rate tendencies in different variants of Mb, L29F < wt < L29A = H64Q < H64F < H64A, and shed light on several aspects of the reaction at the atomic level. First, water access to the distal pocket is a key event and the observed acid catalysis relies on HisE7 protonation and opening of the His gate to allow water access, rather than protonation of the oxy heme itself. Our results also suggest that the basic mechanism, i.e., superoxide displacement by hydroxide anion, is energetically more feasible. Finally, we confirmed that distal hydrogen bonds protect the oxy complex from autoxidation.

High pressure reveals structural determinants for globin hexacoordination: neuroglobin and myoglobin cases.

The influence of pressure on the equilibrium between five-(5c) and six-coordination (6c) forms in neuroglobin (Ngb) and myoglobin (Mb) has been examined by means of molecular dynamics (MD) simulations at normal and high pressure. The results show that the main effect of high pressure is to reduce the protein mobility without altering the structure in a significant manner. Moreover, our data suggest that the equilibrium between 5c and 6c states in globins is largely controlled by the structure and dynamics of the C-D region. Finally, in agreement with the available experimental data, the free energy profiles obtained from steered MD for both proteins indicate that high pressure enhances hexacoordination. In Ngb, the shift in equilibrium is mainly related to an increase in the 6c-->5c transition barrier, whereas in Mb such a shift is primarily due to a destabilization of the 5c state.

Age effects on the social interaction test in early adulthood male rats.

The effects of age on active and passive social interaction were studied in Wistar rats using the social interaction test (S.I.T.). Individual behaviors such as ambulation, rearing, and defecation were also studied. Despite the widespread use of the S.I.T. in anxiety research, the effects of age on the S.I.T. have not been studied thoroughly. Male Wistar rats of 75, 135, and 180 days old were used. Our results showed age effects on active social contact, passive social contact, ambulation, rearing, and defecation. At 135 days old, animals presented the lowest scores on active social behavior and the highest scores on defecation. Moreover, exploratory behavior measured by ambulation and rearing decreased with age. These results suggest that age could be a relevant variable in the social interaction test.