RESUMEN
Hemoglobin (Hb) is a hemeprotein found inside erythrocytes and is crucial in transporting oxygen and carbon dioxide in our bodies. In erythrocytes (Ery), the main energy source is glucose metabolized through glycolysis. However, a fraction of Hb can undergo glycation, in which a free amine group from the protein spontaneously binds to the carbonyl of glucose in the bloodstream, resulting in the formation of glycated hemoglobin (HbA1c), widely used as a marker for diabetes. Glycation leads to structural and conformational changes, compromising the function of proteins, and is intensified in the event of hyperglycemia. The main changes in Hb include structural alterations to the heme group, compromising its main function (oxygen transport). In addition, amyloid aggregates can form, which are strongly related to diabetic complications and neurodegenerative diseases. Therefore, this chapter discusses in vitro protocols for producing glycated Hb, as well as the main techniques and biophysical assays used to assess changes in the protein's structure before and after the glycation process. This more complete understanding of the effects of glycation on Hb is fundamental for understanding the complications associated with hyperglycemia and for developing more effective prevention and treatment strategies.
Asunto(s)
Hemoglobinas , Humanos , Glicosilación , Hemoglobinas/metabolismo , Hemoglobinas/química , Hemoglobina Glucada/metabolismo , Conformación Proteica , AnimalesRESUMEN
Green-stained amniotic fluid, often referred to as meconium-stained amniotic fluid, is present in 5% to 20% of patients in labor and is considered an obstetric hazard. The condition has been attributed to the passage of fetal colonic content (meconium), intraamniotic bleeding with the presence of heme catabolic products, or both. The frequency of green-stained amniotic fluid increases as a function of gestational age, reaching approximately 27% in post-term gestation. Green-stained amniotic fluid during labor has been associated with fetal acidemia (umbilical artery pH <7.00), neonatal respiratory distress, and seizures as well as cerebral palsy. Hypoxia is widely considered a mechanism responsible for fetal defecation and meconium-stained amniotic fluid; however, most fetuses with meconium-stained amniotic fluid do not have fetal acidemia. Intraamniotic infection/inflammation has emerged as an important factor in meconium-stained amniotic fluid in term and preterm gestations, as patients with these conditions have a higher rate of clinical chorioamnionitis and neonatal sepsis. The precise mechanisms linking intraamniotic inflammation to green-stained amniotic fluid have not been determined, but the effects of oxidative stress in heme catabolism have been implicated. Two randomized clinical trials suggest that antibiotic administration decreases the rate of clinical chorioamnionitis in patients with meconium-stained amniotic fluid. A serious complication of meconium-stained amniotic fluid is meconium aspiration syndrome. This condition develops in 5% of cases presenting with meconium-stained amniotic fluid and is a severe complication typical of term newborns. Meconium aspiration syndrome is attributed to the mechanical and chemical effects of aspirated meconium coupled with local and systemic fetal inflammation. Routine naso/oropharyngeal suctioning and tracheal intubation in cases of meconium-stained amniotic fluid have not been shown to be beneficial and are no longer recommended in obstetrical practice. A systematic review of randomized controlled trials suggested that amnioinfusion may decrease the rate of meconium aspiration syndrome. Histologic examination of the fetal membranes for meconium has been invoked in medical legal litigation to time the occurrence of fetal injury. However, inferences have been largely based on the results of in vitro experiments, and extrapolation of such findings to the clinical setting warrants caution. Fetal defecation throughout gestation appears to be a physiologic phenomenon based on ultrasound as well as in observations in animals.
Asunto(s)
Corioamnionitis , Síndrome de Aspiración de Meconio , Complicaciones del Embarazo , Recién Nacido , Embarazo , Femenino , Humanos , Meconio , Líquido Amniótico/química , Inflamación/complicaciones , Hemo/análisisRESUMEN
In this study, we investigated the possibility of interactions between the solvent molecules with the Heme group in the human hemoglobin. The results of this study answer a key question: whether the interactions of the Heme unit with its surroundings are interdependent or independent of the protein units of human hemoglobin. Contributions of the intermolecular interactions were determined by exploiting the solvatochromism spectroscopic data by Kamlet-Taft (KAT) polarity functions. Solvent polarity effects on the nonlinear properties of the Heme's groups in the human hemoglobin (Hb) were investigated via the Z-scan method. The experimental results obtained with spectroscopic and nonlinear optical parameters (absorption coefficient and refractive index) show that the mechanism of solvation and the interactions of Heme are controlled by suitable configuration of the protein units of hemoglobin. In other words, interactions of the Heme with α- and ß-globins are an effective factor in controlling the optical behavior of Heme.
Asunto(s)
Hemo , Hemoglobinas , Humanos , Refractometría , SolventesRESUMEN
Besides multiple life-threatening diseases like lung cancer and cardiovascular disease, cigarette smoking is known to produce hypoxia, a state of inadequate oxygen supply to tissues. Hypoxia plays a pivotal role in the development of chronic obstructive pulmonary disease. Smoking during pregnancy imposes risk for the unborn child. In addition to carbon monoxide, conjugation of para-benzoquinone (pBQ), derived from cigarette smoke, with human hemoglobin (HbA) was also reported to contribute in hypoxia. In fact, conjugation of pBQ is more alarming than carbon monoxide as it is an irreversible covalent modification. In the present study, the functional assay of Hb-pBQ, performed through oxygen equilibrium curve, showed a significant decrease in both P50 and cooperativity. However, the structural changes associated with the observed functional perturbation of the hemoglobin conjugate (Hb-pBQ) are unknown to date. Enhanced sensitivity and high resolution of nano-ESI mass spectrometry platform have enabled to investigate the native structure of oligomers of hemoglobin in a single scan. The structural integrity of Hb-pBQ measured through the dissociation equilibrium constants (Kd) indicated that compared to HbA, Kd of tetramer-dimer and dimer-monomer equilibria were increased by 4.98- and 64.3-folds, respectively. Using isotope exchange mass spectrometry, we observed perturbations in the inter-subunit interactions of deoxy and oxy states of Hb-pBQ. However, the three-dimensional architecture of Hb-pBQ, monitored through collision cross-sectional area, did not show any change. We propose that the significant destabilization of the functionally active structure of hemoglobin upon conjugation with pBQ results in tighter oxygen binding that leads to hypoxia. Graphical Abstract á .
Asunto(s)
Benzoquinonas , Oxígeno , Oxihemoglobinas , Benzoquinonas/química , Benzoquinonas/metabolismo , Medición de Intercambio de Deuterio , Humanos , Espectrometría de Masas , Oxígeno/química , Oxígeno/metabolismo , Oxihemoglobinas/química , Oxihemoglobinas/metabolismo , Unión Proteica , Estabilidad ProteicaRESUMEN
Natural heme proteins may have heme bound to poly-peptide chain as a cofactor via noncovalent forces or heme as a prosthetic group may be covalently bound to the proteins. Nature has used porphyrins in diverse functions like electron transfer, oxidation, reduction, ligand binding, photosynthesis, signaling, etc. by modulating its properties through diverse protein matrices. Synthetic chemists have tried to utilize these molecules in equally diverse industrial and medical applications due to their versatile electro-chemical and optical properties. The heme iron has catalytic activity which can be modulated and enhanced for specific applications by protein matrix around it. Heme proteins can be designed into novel enzymes for sterio specific catalysis ranging from oxidation to reduction. These designed heme-proteins can have applications in industrial catalysis and biosensing. A peptide folds around heme easily due to hydrophobic effect of the large aromatic ring of heme. The directional property of co-ordinate bonding between peptide and metal ion in heme further specifies the structure. Therefore heme proteins can be easily designed for targeted structure and catalytic activity. The central aromatic chemical entity in heme viz. porphyrin is a very ancient molecule. Its presence in the prebiotic soup and in all forms of life suggests that it has played a vital role in the origin and progressive evolution of living organisms. Porphyrin macrocycles are highly conjugated systems composed of four modified pyrrole subunits interconnected at their α -carbon atoms via methine (=CH-) bridges. Initial minimalist models of hemoproteins focused on effect of heme-ligand co-ordinate bonding on chemical reactivity, spectroscopy, electrochemistry and magnetic properties of heme. The great sensitivity of these spectroscopic features of heme to its surrounding makes them extremely useful in structural elucidation of designed heme-peptide complexes. Therefore heme proteins are easier to work on for designing novel proteins for industrial and medical applications.
Asunto(s)
Técnicas Electroquímicas , Hemo/química , Hemoproteínas/química , Porfirinas/química , Ingeniería de Proteínas , Evolución Biológica , Hemoproteínas/síntesis química , Humanos , Interacciones Hidrofóbicas e Hidrofílicas , Ligandos , Modelos Moleculares , Oxidación-Reducción , Pliegue de Proteína , Relación Estructura-ActividadRESUMEN
Pathways of intramolecular conversion and intermolecular electronic excitation energy transfer (EET) in the photosynthetic apparatus of purple bacteria remain subject to debate. Here we experimentally tested the possibility of EET from the bacteriochlorophyll (BChl) Soret band to the singlet S2 level of carotenoids using femtosecond pump-probe measurements and steady-state fluorescence excitation and absorption measurements in the near-ultraviolet and visible spectral ranges. The efficiency of EET from the Soret band of BChl to S2 of the carotenoids in light-harvesting complex LH2 from the purple bacterium Ectothiorhodospira haloalkaliphila appeared not to exceed a few percent.