Effect of vacuum packaging on the biochemical, viscoelastic, and sensory properties of a Spanish cheese during chilled storage

The unique qualities of Spanish cheeses, such as the San Simón da Costa (SSC) cheese, are protected by the Protected Designation of Origin (PDO) status. The technological importance of chilled storage at 4 °C of vacuum-packaged (V) and natural (N) (unpackaged) cheeses was examined. For this purpose, the physico-chemical, biochemical, mechanical (puncture tests), viscoelastic (oscillatory and transient tests) and sensory properties of V and N cheeses were compared and analysed. During chilled storage, the caseins in V cheeses did not undergo proteolytic reactions. Low temperature maintained a low intensity of proteolytic phenomena for up to 6 months. Lipolysis was more intense in the N than in the V samples. The moisture content decreased in the N cheeses during chilled storage, and thus, the casein matrix concentration and ionic strength increased, resulting in an increase in the gel strength (S) parameter and complex modulus (G*), and the conformational stability−high stress amplitude (σmax). The low and similar values of the n’ and n’’ exponents (mechanical spectra) and the n parameter (transient tests) indicated the high degree of the temporal stability of the cheese network in both the N and V samples, irrespective of storage time. Likewise, the similar values of the phase angle (δ) for the N and V cheeses during storage indicate energy-stable bonds in the SSC cheese matrix. The attributes of the oral tactile phase (firmness, friability, gumminess, and microstructure perception), mechanical parameters and viscoelastic moduli enabled the discrimination of the packaged and unpackaged cheeses. Cheeses chilled and stored without packaging were awarded the highest scores for sensory attributes (preference) by trained panellists.Xunta de Galicia | Ref. ED431E 2018/0

The ATPase activity of the DNA transporter TrwB is modulated by protein TrwA: implications for a common assembly mechanism of DNA translocating motors

Conjugative systems contain an essential integral membrane protein involved in DNA transport called the Type IV coupling protein (T4CP). The T4CP of conjugative plasmid R388 is TrwB, a DNA-dependent ATPase. Biochemical and structural data suggest that TrwB uses energy released from ATP hydrolysis to pump DNA through its central channel by a mechanism similar to that used by F1-ATPase or ring helicases. For DNA transport, TrwB couples the relaxosome (a DNA-protein complex) to the secretion channel. In this work we show that TrwA, a tetrameric oriT DNA-binding protein and a component of the R388 relaxosome, stimulates TrwBDeltaN70 ATPase activity, revealing a specific interaction between the two proteins. This interaction occurs via the TrwA C-terminal domain. A 68-kDa complex between TrwBDeltaN70 and TrwA C-terminal domain was observed by gel filtration chromatography, consistent with a 1:1 stoichiometry. Additionally, electron microscopy revealed the formation of oligomeric TrwB complexes in the presence, but not in the absence, of TrwA protein. TrwBDeltaN70 ATPase activity in the presence of TrwA was further enhanced by DNA. Interestingly, maximal ATPase rates were achieved with TrwA and different types of dsDNA substrates. This is consistent with a role of TrwA in facilitating the interaction between TrwB and DNA. Our findings provide a new insight into the mechanism by which TrwB recruits the relaxosome for DNA transport. The process resembles the mechanism used by other DNA-dependent molecular motors, such as the RuvA/RuvB system, to be targeted to the DNA followed by hexamer assembly

Clinical presentation, causative drugs and outcome of patients with autoimmune features in two prospective DILI registries

Background & aims: Idiosyncratic drug-induced liver injury (DILI) with autoimmune features is a liver condition with laboratory and histological characteristics similar to those of idiopathic autoimmune hepatitis (AIH), which despite being increasingly re-ported, remains largely undefined. We aimed to describe in-depth the features of this entity in a large series of patients from two prospective DILI registries. Methods: DILI cases with autoimmune features collected in the Spanish DILI Registry and the Latin American DILI Network were compared with DILI patients without autoimmune features and with an independent cohort of patients with AIH. Results: Out of 1,426 patients with DILI, 33 cases with autoimmune features were identified. Female sex was more frequent in AIH patients than in the other groups (p= .001). DILI cases with autoimmune features had significantly longer time to onset (p< .001) and resolution time (p= .004) than those without autoimmune features. Interestingly, DILI patients with autoimmune features who relapsed exhibited significantly higher total bilirubin and transaminases at onset and absence of peripheral eosinophilia than those who did not relapse. The likelihood of relapse increased over time, from 17% at 6 months to 50% 4 years after biochemical normalization. Statins, nitrofurantoin and minocycline were the drugs most frequently associated with this phenotype. Conclusions: DILI with autoimmune features shows different clinical features than DILI patients lacking characteristics of autoimmunity. Higher transaminases and total bilirubin values with no eosinophilia at presentation increase the likelihood of relapse in DILI with autoimmune features. As the tendency to relapse increases over time, these patients will require long-term follow-up.Instituto de Salud Carlos III; Fondo Europeo de Desarrollo Regional— FEDER, Grant/Award Number: UMA18- FEDERJA-193, PI18/00901, PI19/00883 and PI21/01248; Consejería de Salud y Familia de la Junta de Andalucía, Grant/Award Number: P18-RT- 3364 and PI- 0310- 2018; Agencia Española del Medicamento; Sara Borrell, Grant/Award Number: CD20/00083; Rio Hortega, Grant/Award Number: CM21/00074; Garantía Juvenil, Grant/Award Number: SNGJ5Y6-09; Junta de Andalucía and European Social Fund; European Cooperation in Science and Technology; Universidad de Málaga/CBUA Funding for open access charge: Universidad de Málga / CBU

Substrate translocation involves specific lysine residues of the central channel of the conjugative coupling protein TrwB

Conjugative transfer of plasmid R388 requires the coupling protein TrwB for protein and DNA transport, but their molecular role in transport has not been deciphered. We investigated the role of residues protruding into the central channel of the TrwB hexamer by a mutational analysis. Mutations affecting lysine residues K275, K398, and K421, and residue S441, all facing the internal channel, affected transport of both DNA and the relaxase protein in vivo. The ATPase activity of the purified soluble variants was affected significantly in the presence of accessory protein TrwA or DNA, correlating with their behaviour in vivo. Alteration of residues located at the cytoplasmic or the inner membrane interface resulted in lower activity in vivo and in vitro, while variants affecting residues in the central region of the channel showed increased DNA and protein transfer efficiency and higher ATPase activity, especially in the absence of TrwA. In fact, these variants could catalyze DNA transfer in the absence of TrwA under conditions in which the wild-type system was transfer deficient. Our results suggest that protein and DNA molecules have the same molecular requirements for translocation by Type IV secretion systems, with residues at both ends of the TrwB channel controlling the opening?closing mechanism, while residues embedded in the channel would set the pace for substrate translocation (both protein and DNA) in concert with TrwA

TrwB: Un motor molecular de unión específica a estructuras G-quádruplex en el ADN

La transferencia de ADN por conjugación es un mecanismo de transferencia horizontal que permite la adquisición de nueva información genética de un modo rápido y natural. Este proceso implica un contacto físico entre las células donadora y receptora. Los sistemas conjugativos contienen una proteína clave en la membrana plasmática para llevar a cabo esta función: el transportador de ADN. En nuestro sistema modelo, el plásmido R388, este transportador es la proteína TrwB. La estructura cristalográfica de TrwB¿N70, la fracción soluble de TrwB, revela un hexámero formado por subunidades idénticas y simetría toroidal, con un canal interno de 20 Å de diámetro (Gomis-Ruth et al., 2001). Esta proteína presenta una gran similitud estructural con motores moleculares hexaméricos bien conocidos, tales como el complejo F1-ATPasa (Abrahams et al., 1994), FtsK (Massey et al., 2006) o helicasas en forma de anillo (Singleton et al., 2000), sugiriendo que TrwB también funciona como un motor, utilizando la energía liberada en la hidrólisis de ATP para bombear ADN de cadena sencilla a través de su canal central. En helicasas hexámericas que funcionan en dirección 5'- 3', el extremo 5' pasa a través del canal central de la proteína (Yu et al., 1996) mientras que el extremo 3' contacta la zona externa del hexámero (Ahnert and Patel, 1997). Así pues, de una manera similar, el T-DNA conjugativo sería transferido en dirección 5'-3' a través de la cavidad interna de TrwB. El mecanismo que proponemos para TrwB es similar al que presenta el complejo F1-ATPasa. La hidrólisis secuencial de ATP estaría acoplada a la translocación del ADN a través del poro central, siguiendo un mecanismo similar al >binding change mechanism> seguido por el complejo F1-ATPasa (Abrahams et al., 1994; Boyer, 1993). Sucesivos ciclos de hidrólisis de ATP permitirían cambios conformacionales alternantes en las seis subunidades. Proponemos por lo tanto un mecanismo común que podría aplicarse a una gran variedad de enzimas con un componente estructural común (Cabezon and de la Cruz, 2006).Peer Reviewe

Interactions between TrwB protein and DNA substrates with secondary structures

Trabajo presentado al "35th FEBS Congress: Molecules of Life" celebrado en Gotemburgo (Suecia) del 26 de junio al 1 de julio de 2010.TrwB is a DNA-dependent ATPase involved in DNA transport during bacterial conjugation. The protein presents structural similarity to hexameric molecular motors such as F1-ATPase, FtsK or ring helicases, suggesting that TrwB also operates as a motor, using energy released from ATP hydrolysis to pump ssDNA through its central channel. During this year, we have carried out an extensive analysis with various DNA substrates to determine the preferred substrate for TrwB. Oligonucleotides with G-rich sequences forming G4 DNA structures were the optimal substrates for TrwB ATPase activity. The protein bound with 100-fold higher affinity to G4 DNA than to ssDNA of the same sequence. Moreover, TrwB formed oligomeric protein complexes only with oligonucleotides presenting such a G-quadruplex DNA structure, consistent with stoichiometry of 6 TrwB monomers to G4 DNA, as demonstrated by gel filtration chromatography and analytical ultracentrifugation experiments. G-quadruplex structures are widespread in the genomes and are thought to play a biological function in transcriptional regulation. They form very stable structures that can obstruct DNA replication, requiring the action of specific helicases to resolve them. These observations are discussed in terms of a possible role for TrwB in resolving G4 secondary structures that arise during conjugative DNA processing.Peer Reviewe

Impact of high-pressure processing on antioxidant activity during storage of fruits and fruit products: a review

Fruits and fruit products are an essential part of the human diet. Their health benefits are directly related to their content of valuable bioactive compounds, such as polyphenols, anthocyanins, or vitamins. Heat treatments allow the production of stable and safe products; however, their sensory quality and chemical composition are subject to significant negative changes. The use of emerging non-thermal technologies, such as HPP (High Pressure Processing), has the potential to inactivate the microbial load while exerting minimal effects on the nutritional and organoleptic properties of food products. HPP is an adequate alternative to heat treatments and simultaneously achieves the purposes of preservation and maintenance of freshness characteristics and health benefits of the final products. However, compounds responsible for antioxidant activity can be significantly affected during treatment and storage of HPP-processed products. Therefore, this article reviews the effect of HPP treatment and subsequent storage on the antioxidant activity (oxygen radical absorbance capacity (ORAC) assay), 2,2-diphenyl-1-picrylhydrazyl (DPPH) radical scavenging capacity assay, ferric reducing antioxidant power (FRAP) assay, 2,2′-azino-bis-(3-ethylbenzothiazoline-6-sulfonic acid) (ABTS) radical scavenging capacity assay or Trolox equivalent antioxidant capacity (TEAC) assay), and on the total phenolic, flavonoid, carotenoid, anthocyanin and vitamin contents of fruits and different processed fruit-based products

Amino acid profile and protein quality related to canning and storage of swordfish packed in different filling media

Financiado para publicación en acceso aberto: Universidade de Vigo/CISUGThis study was conducted to determine the effect of canning, different filling media (olive oil, corn oil, sunflower oil and high oleic sunflower oil) and storage on the amino acid profile and protein quality in swordfish. Glutamic acid, aspartic acid and leucine are the predominant amino acids in raw swordfish, representing 35.62 % of the total amino acids. Thawing-salting caused significant losses of methionine, glycine and tyrosine. Frying caused moisture loss, resulting in higher protein and amino acid contents in fried swordfish than in raw or salted fish. The changes observed during the sterilization process depended on the amino acid and filling medium. The concentrations of some amino acids, such as methionine and glycine, decreased during heat treatment; however, the proline content increased. Slight changes in some amino acids were detected after storage of the canned swordfish for 12 months. The changes during storage were also influenced by the filling medium and varied depending on the amino acid considered. The highest scores in raw and canned swordfish were observed for histidine. The quality indices used showed that canned swordfish packed in olive oil was the highest quality product of those considered.Ministerio de Medio Ambiente, y Medio Rural y Marino | Ref. ARM/1790/201

Free amino acids and biogenic amines in canned European fels: influence of processing step, filling medium and storage time

This study evaluated the effects of the canning process and different filling media on the free amino acid and biogenic amine contents of eels. The main free amino acids were histidine, taurine and arginine, which constituted 72% of the free amino acids in raw eels. All steps in the canning process significantly altered the free amino acid content of eels, relative to raw samples. The changes were influenced by the step, the composition of the frying or filling medium and the storage time. The biogenic amine contents were very low in all samples. Histamine was not detected in either raw eels or canned eels. The highest values were obtained for 2-phenylethylamine. The step of the canning process, the composition of the frying or filling medium and storage time also determined the changes in the biogenic amine contents. The biogenic amines indices were low, indicating the good quality of canned eels

Structural and Theoretical Evidence of the Depleted Proton Affinity of the N3-Atom in Acyclovir

The hydronium salt (H3O)2[Cu(N7–acv)2(H2O)2(SO4)2]·2H2O (1, acv = acyclovir) has been synthesized and characterized by single-crystal X-ray diffraction and spectral methods. Solvated Cu(OH)2 is a by-product of the synthesis. In the all-trans centrosymmetric complex anion, (a) the Cu(II) atom exhibits an elongated octahedral coordination; (b) the metal-binding pattern of acyclovir (acv) consists of a Cu–N7(acv) bond plus an (aqua)O–H···O6(acv) interligand interaction; and (c) trans-apical/distal sites are occupied by monodentate O-sulfate donor anions. Neutral acyclovir and aqua-proximal ligands occupy the basal positions, stabilizing the metal binding pattern of acv. Each hydronium(1+) ion builds three H-bonds with O–sulfate, O6(acv), and O–alcohol(acv) from three neighboring complex anions. No O atoms of solvent water molecules are involved as acceptors. Theoretical calculations of molecular electrostatic potential surfaces and atomic charges also support that the O-alcohol of the N9(acv) side chain is a better H-acceptor than the N3 or the O-ether atoms of acv