Spectral characterisation of red pigment in Italian-type dry-cured ham. Increasing lipophilicity druing processing and maturation

Spectroscopic studies of Parma ham during processing revealed a gradual transformation of muscle myoglobin, initiated by salting and continuing during ageing. Electron spin resonance spectra did, however, conclusively show that the pigment in dry-cured Parma ham at no stage is a nitrosyl complex of ferrous myoglobin as found in brine-cured ham and Spanish Serrano hams. Both near-infra red reflectance spectra of sliced ham and UV/visible absorption spectra of extract of hams, obtained with aqueous buffer or acetone, showed the presence of different red pigments at varying processing stages for both solvents. Especially, the pigment extracted with aqueous buffer exhibited unique spectral features different from those of well-known myoglobin derivatives. At the end of processing, the pigment(s) becomes less water extractable, while the fraction of red pigment(s) extractable with acetone/water (75%/25%) increases throughout the processing time up to full maturation at 18 months. The chemical identity of the 6th ligand of myoglobin could not be conclusively established, but possible candidates are discussed. The partition of the pigment(s) between pentane and acetone/water showed a strong preference for pentane, suggesting that only the heme moiety is present in the acetone/water extract, and that Parma ham pigment is gradually transformed from a myoglobin derivative into a non-protein heme complex, which was found to be thermally stable in acetone/water solutio

Oxidation of Native and Modified Hemoglobin and Myoglobin by Sodium Nitrate. Effect of Inositol Hexaphosphate

Native and modified hemoglobin, myoglobin and a and phemoglobin subunits were oxidized by sodium nitrite at pH 6. The experiments were carried out under oxy and deoxy conditions with and without inositol hexaphosphate (IHP). It is shown (a) that under oxy condition low concentration of IHP inhibits the oxidation of native hemoglobin only. However, high concentration of IHP inhibits the oxidation of both myoglobin and modified hemoglobin (digested or 0-93-SH groups blocked). This inhibition is partially counteracted by high oxygen pressure, (b) Under deoxy condition the oxidation rates of all hemeproteins studied are significantly faster than that of native hemoglobin. IHP inhibits the oxidation of all except the myoglobin and hemoglobin subunits. It is concluded that although the IHP inhibitory effect on hemoglobin oxidation by nitrite can be explained by the shift of the R↔T conformational equilibrium towards T conformation, some other structural changes such as alteration in molecular surface charges must occur to account for the effect of IHP on the oxidation of hemeproteins devoid of heme-heme interaction

An investigation into the feasibility of myoglobin-based single-electron transistors

Myoglobin single-electron transistors were investigated using nanometer- gap platinum electrodes fabricated by electromigration at cryogenic temperatures. Apomyoglobin (myoglobin without heme group) was used as a reference. The results suggest single electron transport is mediated by resonant tunneling with the electronic and vibrational levels of the heme group in a single protein. They also represent a proof-of-principle that proteins with redox centers across nanometer-gap electrodes can be utilized to fabricate single-electron transistors. The protein orientation and conformation may significantly affect the conductance of these devices. Future improvements in device reproducibility and yield will require control of these factors

Solvent-induced organization: A physical model of folding myoglobin

The essential features of the in vitro refolding of myoglobin are expressed in a solvable physical model. Alpha helices are taken as the fundamental collective coordinates of the system, while the refolding is assumed to be mainly driven by solvent-induced hydrophobic forces. A quantitative model of these forces is developed and compared with experimental and theoretical results. The model is then tested by being employed in a simulation scheme designed to mimic solvent effects. Realistic dynamic trajectories of myoglobin are shown as it folds from an extended conformation to a close approximation of the native state. Various suggestive features of the process are discussed. The tenets of the model are further tested by folding the single-chain plant protein leghemoglobin.Comment: Rockefeller preprint RU 93-3-B 28 pages, plain LATEX Figures available by request to [email protected]

Label-Free, Highly Sensitive Electrochemical Aptasensors Using Polymer-Modified Reduced Graphene Oxide for Cardiac Biomarker Detection

Acute myocardial infarction (AMI), also recognized as a ???heart attack,??? is one leading cause of death globally, and cardiac myoglobin (cMb), an important cardiac biomarker, is used for the early assessment of AMI. This paper presents an ultrasensitive, label-free electrochemical aptamer-based sensor (aptasensor) for cMb detection using polyethylenimine (PEI)-functionalized reduced graphene oxide (PEI???rGO) thin films. PEI, a cationic polymer, was used as a reducing agent for graphene oxide (GO), providing highly positive charges on the rGO surface and allowing direct immobilization of negatively charged single-strand DNA aptamers against cMb via electrostatic interaction without any linker or coupling chemistry. The presence of cMb was detected on Mb aptamer-modified electrodes using differential pulse voltammetry via measuring the current change due to the direct electron transfer between the electrodes and cMb proteins (Fe3+/Fe2+). The limits of detection were 0.97 pg mL???1 (phosphate-buffered saline) and 2.1 pg mL???1 (10-fold-diluted human serum), with a linear behavior with logarithmic cMb concentration. The specificity and reproducibility of the aptasensors were also examined. This electrochemical aptasensor using polymer-modified rGO shows potential for the early assessment of cMb in point-of-care testing applications

Review of centrifugal liquid-liquid chromatography using aqueous two-phase solvent (ATPS) systems: Its scale-up and prospects for the future production of high-value biologics

The future challenges in bioprocessing include developing new downstream processes for the purification and manufacture of the protein based medicines of the future to relieve the predicted bottleneck being produced by increasingly high titres from fermentation processes. This review looks at the recent developments in centrifugal liquid-liquid partition chromatography using aqueous two-phase solvent (ATPS) systems, a gentle host medium for biologics, and the prospect for scale-up and eventual manufacture of high value pharmaceutical products

Femtosecond X-ray emission study of the spin cross-over dynamics in haem proteins

In haemoglobin (consisting of four globular myoglobin-like subunits), the change from the low-spin (LS) hexacoordinated haem to the high spin (HS) pentacoordinated domed form upon ligand detachment and the reverse process upon ligand binding, represent the transition states that ultimately drive the respiratory function. Visible-ultraviolet light has long been used to mimic the ligand release from the haem by photodissociation, while its recombination was monitored using time-resolved infrared to ultraviolet spectroscopic tools. However, these are neither element- nor spin-sensitive. Here we investigate the transition state in the case of Myoglobin-NO (MbNO) using femtosecond Fe Kalpha and Kbeta non-resonant X-ray emission spectroscopy (XES) at an X-ray free-electron laser upon photolysis of the Fe-NO bond. We find that the photoinduced change from the LS (S = 1/2) MbNO to the HS (S = 2) deoxy-myoglobin (deoxyMb) haem occurs in ca. 800 fs, and that it proceeds via an intermediate (S = 1) spin state. The XES observables also show that upon NO recombination to deoxyMb, the return to the planar MbNO ground state is an electronic relaxation from HS to LS taking place in ca. 30 ps. Thus, the entire ligand dissociation-recombination cycle in MbNO is a spin cross-over followed by a reverse spin cross-over process