Prokaryotic expression and functional analysis of molecular chaperone Acr2 protein of Mycobacterium tuberculosis / 中国生物制品学杂志

@#Objective To express the molecular chaperone Acr2 protein of Mycobacterium tuberculosis(Mtb)in E.coli and analyze the function. Methods The recombinant plasmid pET-28a-Acr2 was transformed into competent E. coli BL21(DE3),and induced by IPTG. The expressed His-Acr2 protein was purified by Ni-NTA chromatography and SuperdexTM200 10/300 GL gel filtration chromatography to obtain Acr2 protein. The Acr2 protein was refolded by spontaneous refolding and reassembly after thermal denaturation(100 ℃ for 15 min)and chemical denaturation(8 mol/L urea,37 ℃ for 4 h).The secondary structure of Acr2 protein before and after denaturation-renaturation was detected by circular dichroism spectroscopy and non-denaturing SDS-PAGE,and the molecular chaperone function of Acr2 protein in vitro was detected by substrate binding assay. Results The purified Acr2 protein had the relative molecular mass of about 232 000,the purity of over 90%,and the concentration of about 2 mg/mL,which recovered its natural secondary structure after denaturationrenaturation,and formed stable complexes with the denatured malate dehydrogenase(MDH)at 48 ℃. Conclusion The Acr2protein can restore its natural molecular conformation with molecular chaperone activity in vitro after denaturation-renaturation treatment,providing a new strategy for the preparation of Mtb protein antigen with natural activity.

Optimization of expression and purification conditions of recombinant Hq001 protein in salivary glands of Haemaphysalis qinghaiensis / 中国生物制品学杂志

@#Objective To optimize the expression conditions(expression and induction conditions)and purification methods(non-denaturing and denaturing purification)of recombinant Hq001 protein in salivary glands of Haemaphysalis qinghaiensis.Methods The recombinant plasmid pET-30a-Hq001 was transformed into competent cells E.coil BL21(DE3),E.coil Rosetta(DE3)and E.coil ArcticExpress(DE3)pRARE2 respectively for the selection of an optimal expression strain.The final concentration of IPTG(0,0.5,1.0 mmol/L),induction temperature(20,25 ℃)and induction time(0,2,4,6,8 h)were optimized.The recombinant bacteria expressed under the ideal induction condition were homogenized by French press and the target protein was purified by passing through a Ni-NTA affinity chromatography column under either native(denaturationrenaturation-column chromatography)or denatured conditions(denaturation-column chromatography-renaturation).The purified products were analyzed by 12% SDS-PAGE.Results E.coil BL21(DE3)was proved to be the most suitable strain for the expression of recombinant Hq001 protein.The optimum induction condition was induction with 0.5 mmol/L IPTG for 4 h at 25 ℃.The target protein with a relative molecular mass of approximately 18 800 was obtained by non-denaturing purification method,and the size was consistent with the expectation.Conclusion The recombinant protein rHq001 in salivary glands of Haemaphysalis qinghaiensis can be obtained by the optimized expression conditions and purification methods.

Reducción de proteínas y glucosa por reacción de Maillard en leche con lactosa hidrolisada / Protein and glucose reduction by Maillard reaction in milk with hydrolyzed lactose

Se evaluó el efecto de la temperatura sobre la desnaturalización de proteínas y la reacción de Maillard en leche entera y descremada con lactosa hidrolizada. Las leches hidrolizadas se trataron térmicamente a 100, 110, 120 y 130 °C durante un período de 1 hora y se midió la concentración de glucosa, el grado de pardeamiento y la desnaturalización de proteínas. El grado de dorado en la leche entera varió de 14.4 (100 °C) a 42.6 (130 °C). Para la leche descremada fue de 20.2 (100 °C) a 38.0 (130 °C). La concentración de glucosa en leche entera (47% p/v) y en leche descremada (41% p/v) después del tratamiento térmico (130 °C) mostró una reducción significativa en relación con el control (25 °C). El efecto de la temperatura en la desnaturalización de proteínas en leche entera y descremada en relación con el control (25 °C) fue del 100%. La leche tratada térmicamente con lactosa hidrolizada promovió la desnaturalización de proteínas con un aumento del pardeamiento característico de la reacción de Maillard, lo que afectó la calidad nutricional.

The effect of temperature in protein denaturation and Maillard reaction in whole and skim milk with hydrolyzed lactose was evaluated. Hydrolyzed milk was thermally treated at 100, 110, 120 and 130 °C over a period of 1 hour and glucose concentration, browning degree and protein denaturation were measured. The browning degree in whole milk varied from 14.42 (100 °C) to 42.63 (130 °C) and 20.21 (100 °C) to 38.03 (130 °C) in skim milk. Glucose concentration in whole milk (47% - w/v) and skim milk (41% - w/v) after heat treatment (130 °C) showed a significant reduction in relation to the control (25 °C). The temperature effect in protein denaturation in whole and skim milk in relation to the control (25 °C) was 100%. Thermally treated milk with hydrolyzed lactose promoted protein denaturation with increasing browning characteristic of the Maillard reaction, thus affecting the nutritional quality.

In vitro biological activities of aqueous extracts of Tetrapleura tetraptera (Schumach. & Thonn.) taub. and Aframomum citratum (C. Pereira) K.Schum from three Agroecologic Zones in Cameroon

Objective: To investigate some phytochemical constituents and biological activities of twelve samples of Tetrapleura tetraptera (Schumach & Thonn.) taub. and nine samples of Aframomum citratum (C. Pereira) K. Schum fruits collected in the bimodal forest zone (ZONE V), the unimodal forest zone (ZONE IV) and the highlands zone (ZONE III) in Cameroon. Methods: Fresh fruits extracts were obtained by aqueous infusion (100 °C during 15 min) and evaluated by spectrophotometric methods for total polyphenol (TPP), total flavonoids (TFLV) contents and antioxidant (DPPH, total antioxidant capacity by the phosphomolybdenum method, iron reducing power or ferric reducing antioxidant power and inhibition of beta carotene discoloration assays) and anti-inflammatory (inhibitions of protein denaturation and 5-LOX represented by INH.PROT and INH.5- LOX respectively) properties. Principal component analysis was performed. Results: For both species, fruits from ZONE V have the highest TPP, TFLV levels and biological activities. TPP and TFLV content of Aframomum citratum and Tetrapleura tetraptera fruits are positively and significantly (P<0.05) correlated. The biological activities of all extracts (0.25, 2.5, 25, 250 mg/mL) were dosedependent and the extracts have shown strong antioxidant and anti- inflammatory activities, but less than references (ascorbic acid, diclofenac, quercetin, and butylated hydroxytoluene). There was a positive correlation between TPP, TFLV and total antioxidant capacity, ferric reducing antioxidant power, and inhibition of beta carotene discoloration assays, and inverse correlations were observed with the IC50 (g/mL) of DPPH, INH.5-LOX and INH. PROT assays for both species. Conclusions: The fruits exhibit variabilities and those from ZONE V for both species are economically and healthcare challenging for herbalists, pharmaceutical firms, scientists and consumers. Indeed, most important extraction yield of bioactive compounds correlated with significant biological activities and the use of less material compared with an implementation in other Agro-ecologic Zones with the same results are noted.

In vitro biological activities of aqueous extracts of Tetrapleura tetraptera (Schumach. & Thonn.) taub. and Aframomum citratum (C. Pereira) K.Schum from three Agroecologic Zones in Cameroon

Objective: To investigate some phytochemical constituents and biological activities of twelve samples of Tetrapleura tetraptera (Schumach & Thonn.) taub. and nine samples of Aframomum citratum (C. Pereira) K. Schum fruits collected in the bimodal forest zone (ZONE V), the unimodal forest zone (ZONE IV) and the highlands zone (ZONE III) in Cameroon. Methods: Fresh fruits extracts were obtained by aqueous infusion (100 °C during 15 min) and evaluated by spectrophotometric methods for total polyphenol (TPP), total flavonoids (TFLV) contents and antioxidant (DPPH, total antioxidant capacity by the phosphomolybdenum method, iron reducing power or ferric reducing antioxidant power and inhibition of beta carotene discoloration assays) and anti-inflammatory (inhibitions of protein denaturation and 5-LOX represented by INH.PROT and INH.5- LOX respectively) properties. Principal component analysis was performed. Results: For both species, fruits from ZONE V have the highest TPP, TFLV levels and biological activities. TPP and TFLV content of Aframomum citratum and Tetrapleura tetraptera fruits are positively and significantly (P<0.05) correlated. The biological activities of all extracts (0.25, 2.5, 25, 250 mg/mL) were dosedependent and the extracts have shown strong antioxidant and anti- inflammatory activities, but less than references (ascorbic acid, diclofenac, quercetin, and butylated hydroxytoluene). There was a positive correlation between TPP, TFLV and total antioxidant capacity, ferric reducing antioxidant power, and inhibition of beta carotene discoloration assays, and inverse correlations were observed with the IC

In Vitro Anti-Inflammatory Activity Of Syringic Acid

Objective: The present study was carried out to investigate the in vitro anti-inflammatory activity of syringic acid (SA).  Methods: SA was tested for it's in vitro anti-inflammatory activity at different concentrations in protein denaturation, proteinase inhibition and human red blood cell (HRBC) membrane stabilization assay. The reference drugs used were aspirin and diclofenac sodium. Results: SA showed concentration-dependent inhibition of protein denaturation and proteinase activity with a half-maximal inhibitory concentration (IC50) value of 49.38±0.56 µg/ml and 53.73±0.27 µg/ml respectively. Heat-induced haemolysis was inhibited by SA with an IC50 value of 57.13±0.24 µg/ml. SA also inhibited the hypotonicity-induced haemolysis (IC50 value of 53.87±0.72 µg/ml). Conclusion: From the present study, we can conclude that SA possesses appreciable anti-inflammatory effect against denaturation of proteins, proteinase activity, and human red blood membrane stabilization assays. Further studies are required for determining the possible mechanisms behind its anti-inflammatory action.

Evaluation of in vitro and in vivo therapeutic efficacy of Ribes alpestre Decne in Rheumatoid arthritis

Rheumatoid arthritis is an autoimmune inflammatory disorder, despite the discovery of numerous drugs there is still need to introduce newer, safer and more effective sources of drugs such as medicinal herbs. Present research work was an attempt to appraise the antiarthritic potential of Ribes alpestre Decne in rheumatoid arthritis. In vitro inhibition of protein (bovine serum albumin and egg albumin) denaturation, Human red blood cell membrane stabilization assays along with formaldehyde induced arthritis in rats were commenced in this study. Findings of present investigation demonstrated significant and dose dependent antiarthritic effect of Ribes alpestre. Aqueous ethanolic extract, butanol and aqueous fraction illustrated 95%, 69.233% and 92.840% protection at 6400 ug/mL against bovine serum albumin denaturation respectively. Similarly, plant extract together with butanol and aqueous fractions showed 3653.47%, 1484.03% and 3563.19% inhibition of pathological alteration of egg albumin in that order. Moreover, hydroethanolic extract with butanol and aqueous fraction exhibited 91.29%, 65.73% and 89.62% stabilization against erythrocyte hemolysis at 6400 ug/mL correspondingly. Furthermore, hydroethanolic extract, butanol and aqueous fraction notably 73.49%, 66.42% and 68.87% decreased paw edema at highest dose (200 mg/kg). Similarly aqueous ethanolic extract, butanol and aqueous fraction illustrated 72.38%, 54.90% and 66.33% decrease in paw thickness at 200 mg/kg. Hence results suggested that Ribes alpestre possess antiarthritic potential thus supporting its use as natural remedy in rheumatic conditions.

In Vitro Anti-arthritic Activity Of Kashaya Of Sida Cordifolia Linn.

Arthritis is a form of joint disorder that involves inflammation of one or more joints. It is very common condition especially in women and older people. Ayurveda has contributed a lot for the management of arthritic conditions. The plant Bala, identified as Sida cordifolia Linn. is a widely used drug in many of the Ayurvedic formulations especially in those for arthritic conditions. Kashaya (decoction) is one of the commonly prescribed preparations in Ayurveda. Present study was aimed to assess the anti-arthritic activity of Kashaya (decoction) of root of Sida cordifolia Linn. by inhibition of protein denaturation method and Inhibition of proteinase enzyme activity. Kashaya of roots of Bala was prepared as per standard procedure and was used to induce protein denaturation in Bovine serum albumin and to inhibit the activity of proteinase enzyme, trypsin. The absorbance was read by spectrophotometer to evaluate the percentage of inhibition in both the procedures. Each experiment was done in triplicates. The results were compared with standard drug Diclofenac sodium. Sida cordifolia Linn. showed dose dependent inhibitory activity and highest activity was seen in 500µg/ml concentration in both the experiments. The result showed that root of Sida cordifolia Linn. is having anti-arthritic property. Further studies can be carried out with other formulations of Bala like Choorna (powder), Swarasa (juice) etc. to compare their anti-arthritic activity. The study supports the classical use of plant Bala in various formulations in the treatment of arthritis.

Denaturation studies on bovine serum albumin–bile salt system:Bile salt stabilizes bovine serum albumin through hydrophobicity / 药物分析学报

Protein denaturation is under intensive research, since it leads to neurological disorders of severe con-sequences. Avoiding denaturation and stabilizing the proteins in their native state is of great importance, especially when proteins are used as drug molecules or vaccines. It is preferred to add pharmaceutical excipients in protein formulations to avoid denaturation and thereby stabilize them. The present study aimed at using bile salts (BSs), a group of well-known drug delivery systems, for stabilization of proteins. Bovine serum albumin (BSA) was taken as the model protein, whose association with two BSs, namely sodium cholate (NaC) and sodium deoxycholate (NaDC), was studied. Denaturation studies on the pre-formed BSA-BS systems were carried out under chemical and physical denaturation conditions. Urea was used as the chemical denaturant and BSA-BS systems were subjected to various temperature conditions to understand the thermal (physical) denaturation. With the denaturation conditions prescribed here, the data obtained is informative on the association of BSA-BS systems to be hydrophobic and this effect of hydrophobicity plays an important role in stabilizing the serum albumin in its native state under both chemical and thermal denaturation.

Dose-effect relationships of bipolar radio frequency heating / 中华物理医学与康复杂志

Objective To study the effect of the preset temperature,heating time and the spacing of radio frequency electrodes.Methods Fresh egg white was used to study the effects of heater spacing,temperature and heating time.Two electrodes were fixed to keep the needles perpendicular to the center of egg white confined on a square plate.Temperatures from 50 to 90℃ were tested with heating times of 60 and 120 seconds and electrode spac ings of 2.0,1.0,0.5 and 0.25 centimetres.The real-time temperature and the time to the appearance of a spindleshaped bridge between the electrodes was observed.The egg white's impedance and resistance,the lowest temperature of denaturation,and the mass (or volume) denatured at different time points were also observed.Results With an electrode spacing of 2 cm and the thermostat set at 63℃,the true electrode temperatures were 63.7 and 52.6℃,with the secondary electrode the cooler.Significant differences between the secondary electrode temperature and the preset temperature as well as the temperature between the primary and the secondary electrodes were observed.It was also found that different electrode spacing caused significant differences between the preset temperature and the primary as well as the secondary electrode temperature.Denaturation began at 56℃ with a spacing of 0.25 cm.At spacings of 0.25 and 0.50 cm,a spindle-shaped connection between the two electrodes was observed at temperatures above 70℃,but higher temperature was required at 1.00 and 2.00 cm.When the spacing was 0.25 cm and the preset temperature was 75℃,the denatured volume after 60 s of heating was 90.21+0.64 mm3.Heating for 120 s denatured 95.08+ 0.53 mm3 and two 60.s heatings 92.88+0.74 mm3,all significant differences.There were no significant differences in the resistance of the egg white before and after a single heating for 60 s,but after a single heating lasting 120 s it had increased significantly from 128.41+8.04 to 121.29±8.04 Ω.Conclusion Bipolar radio frequency heating can denature egg white.Higher temperature,longer heating and smaller electrode spacing heat more effectively.

Kinetics and Thermodynamics of Thermal Inactivation of β-Galactosidase from Aspergillus oryzae

ABSTRACT For optimization of biochemical processes in food and pharmaceutical industries, the evaluation of enzyme inactivation kinetic models is necessary to allow their adequate use. Kinetic studies of thermal inactivation of β-galactosidase from Aspergillus oryzae were conducted in order to critically evaluate mathematical equations presented in the literature. Statistical analysis showed that Weibull model presented the best adequacy to residual enzymatic activity data through the processing time and its kinetic parameters as a function of the temperature, in the range of 58-66 ºC. The investigation suggests the existence of a non-sensitive heat fraction on the enzyme structure, which is relatively stable up to temperatures close to 59 ºC. Thermodynamic parameters were evaluated and showed that such β-galactosidase presents activation energy of 277 kJ mol-1 and that the enzyme inactivation is due to molecular structural changes. Results shown that the enzyme is quite stable for biotechnological applications.

Characterization of denaturation and renaturation of DNA for DNA hybridization

OBJECTIVES: The present study was designed to systematically characterize the denaturation and the renaturation of double stranded DNA (dsDNA), which is suitable for DNA hybridization. METHODS: A series of physical and chemical denaturation methods were implemented on well-defined 86-bp dsDNA fragment. The degree of each denaturation was measured and the most suitable denaturation method was determined. DNA renaturation tendency was also investigated for the suggested denaturation method. RESULTS: Heating, beads mill, and sonication bath did not show any denaturation for 30 minutes. However probe sonication fully denatured DNA in 5 minutes. 1 mol/L sodium hydroxide (alkaline treatment) and 60% dimethyl sulfoxide (DMSO) treatment fully denatured DNA in 2-5 minutes. CONCLUSIONS: Among all the physical methods applied, the direct probe sonication was the most effective way to denature the DNA fragments. Among chemical methods, 60% DMSO was the most adequate denaturation method since it does not cause full renaturation during DNA hybridization.

Prognostic value of multiple-gene joint detection in bladder cancer / 中国肿瘤临床

Objective: To investigate the molecular changes in bladder urothelial carcinoma via different pathways. Methods:Polymerase-chain reaction (PCR) or coamplification at low denaturation temperature-PCR and Sanger direct sequencing were per-formed to detect the status of fgfr3, p53, and h-ras gene mutations in 88 tissue samples of human bladder cancer and 10 normal control tissues. The relative mRNA expression levels of motility-related protein-1 (MRP-1)/CD9 and the relationship between genes and tumor recurrence were also determined. Logistic regression and relative analyses were conducted to compare the significance and interrelation of genes among tumor recurrences. Results:The mutation rate of p53 increased as pathological grades and stages increased. Recurrence rate was higher in patients with MT-p53 genotype than in patients with WT-p53 genotype. Conversely, the mutation rate of fgfr3 gene decreased as pathological grades and stages increased. Recurrence rate was also higher in patients with WT-fgfr3 genotype than in pa-tients with MT-fgfr3 genotype. In low-grade and early stage tumors, MT-fgfr3/WT-p53 was the most prevalent genotype;in high-grade and late stage tumors, WT-fgfr3/MT-p53 was the most prevalent genotype. The mutations of h-ras were mainly observed in low-grade tumors in early stages. Moreover, the relative mRNA levels of MRP-1/CD9 decreased as pathological grades and stages increased. The mRNA levels of MRP-1/CD9 were negatively correlated with p53 mutations and positively correlated with fgfr3 mutations. Logistic re-gression analysis results showed that patients with WT-fgfr3 genotypes exhibited 3.88 times higher relative risk of tumor recurrence than those with MT-fgfr3 genotypes;by contrast, patients with MT-p53 genotypes exhibited 4.53 times higher relative risk of tumor re-currence than those with WT-p53 genotypes. Conclusion:Fgfr3 and h-ras gene mutations may play important roles in tumorigenesis of low-grade and early stage bladder cancer. p53 gene mutation and mRNA levels of MRP-1/CD9 may be implicated in the tumorigenesis of high-grade tumors in late stage of bladder cancer. In general, the two variants of urothelial carcinoma exhibit distinct genetic defects. fgfr3 gene mutation revealed a pathway of favorable prognosis, and p53 gene mutation demonstrated a pathway associated with poor prognosis.

Residual ordered structure in denatured proteins and the problem of protein folding.

Structural characteristics of numerous globular proteins in the denatured state have been reviewed using literature data. Recent more precise experiments show that in contrast to the conventional standpoint, proteins under strongly denaturing conditions do not unfold completely and adopt a random coil state, but contain significant residual ordered structure. These results cast doubt on the basis of the conventional approach representing the process of protein folding as a spontaneous transition of a polypeptide chain from the random coil state to the unique globular structure. The denaturation of proteins is explained in terms of the physical properties of proteins such as stability, conformational change, elasticity, irreversible denaturation, etc. The spontaneous renaturation of some denatured proteins most probably is merely the manifestation of the physical properties (e.g., the elasticity) of the proteins per se, caused by the residual structure present in the denatured state. The pieces of the ordered structure might be the centers of the initiation of renaturation, where the restoration of the initial native conformation of denatured proteins begins. Studies on the denaturation of proteins hardly clarify how the proteins fold into the native conformation during the successive residue-by-residue elongation of the polypeptide chain on the ribosome.

Further studies on membrane stabilizing, anti-inflammatory and FCA induced arthritic activity of various fractions of bark of Machilus macrantha in rats

Machilus macrantha Nees, Lauraceae, bark is traditionally used in the treatment of asthma, tuberculosis and rheumatoid arthritis. In order to validate, mechanism based anti-inflammatory activity of fractions M. macrantha bark are investigated for first time. Test materials viz. petroleum ether (PE), alkaloidal fraction (CH), acetone extracts (TAN) and mucilage (MM) (250 and 500 mg/kg, p.o.) obtained from M. macrantha bark were tested for membrane stabilizing, anti-nociceptive; anti inflammatory and Freund's complete adjuvant (FCA) induced arthritis activity. Diclofenac sodium and morphine were used as the reference standards in pharmacological assay. Test materials have significantly (p<0.01) inhibited paw edema after Carrageenan and histamine induction at higher doses. Administration of test materials of M. macrantha (250 and 500 mg/kg b.w.) significantly reduced abdominal writhing, formalin nociception, cotton pellet granuloma and vascular permeability in experimental animal. In addition to this, bark of M. macrantha showed chronic anti-rheumatic effect by suppressing the swelling volume, arthritis index, hematological and biochemical parameters (ESR, RA factor, CRP, liver transferase enzyme) in FCA-induced arthritis. It also significantly inhibited protein denaturation, heat-induced haemolysis of RBC and reduction in total leukocyte migration. Bioassay guided fractionation of the pet. ether extract of bark of M. macrantha led to isolation and characterization of β-sitosterol and stigma sterol confirmed by its HPLC, NMR and GC-MS study. In conclusion, extracts of M. macrantha bark can be explored as a therapeutic agent for the treatment of acute and chronic arthritis.

Hemoglobin F level in different hemoglobin variants

BACKGROUND: Fetal hemoglobin (HbF) levels in different hemoglobin variants in Osogbo, Nigeria, were estimated using two principal methods of estimation using existing information for HbF concentration and distribution of various hemoglobin variants in the area, as well as diagnosed cases of thalassemia. Two hundred and sixty samples collected from HbSS, HbSC, HbAA, HbAS, and HbAC subjects were analyzed. HbF level and hemoglobin type were determined in this study. METHODS: The hemoglobin type was determined using cellulose acetate electrophoresis at an alka-line pH, and HbF was determined by the acid elution and alkaline denaturation methods. RESULTS: The mean+/-SD of HbF in the respective hemoglobin variants was as follows: HbSS, 2.09+/-1.94%; HbSC, 0.85+/-0.54%; HbAA, 0.69+/-0.46%; HbAS, 0.52+/-0.31%; and HbAC, 0.57+/-0.26%. The mean HbF level across the hemoglobin variants was statistically significant (P<0.05). Investigating the sex distribution of the HbF level in the studied population revealed a slightly higher mean HbF level in females than in their male counterparts. CONCLUSION: Within the study population, the HbF level was found to be highest in HbSS and lowest in HbAS. The two methods of estimating HbF are equally reliable, since there was no significant difference between the results obtained from the two methods.

ESTABILIZACIÓN DE HOLO-α-LACTOALBÚMINA EN PRESENCIA DE POLIOLES / HOLO-α-LACTALBUMLN STABILIZATION IN THE PRESENCE OF POLYOLS / ESTABILIDADE TÉRMICA DA HOLO-α-LACTOALBUMFNA EM PRESENÇA DE POLIÓIS

En este trabajo se presenta un estudio sistemático sobre el efecto de las soluciones acuosas de eritritol, xilitol, sorbitol e inositol con diferentes concentraciones, sobre la estabilidad térmica de la holo-α-lactoalbúmina bovina con pH 6,5 usando espectroscopia UV-VIS. Los resultados obtenidos muestran que los polioles usados estabilizan la holo-α-lactoalbúmina en un grado significativamente menor al reportado para otras proteínas. Se sugiere que este menor efecto de estabilización ocurre debido a que esta proteína presenta un estado desnaturalizado parcialmente desdoblado.

In this work we present a systematic study of the effect of aqueous solutions of erythritol, xylitol, sorbitol and inositol on thermal stability of bovine holo-α-lactalbumin at pH 6,5 using UV-VIS spectroscopy. The results show that the polyols used stabilize the holo-α-lactoalbumin in a significant lesser extent than the reported for others proteins. It is suggested that the lower stabilization achieved for this protein is the result of a partially unfolded denaturated state that this protein presents.

Neste trabalho é apresentado um estudo sistemático acerca do efeito de soluções aquosas de eritritol, xilitol, sorbitol e inositol a diferentes concentrações sobre a estabilidade térmica da holo-α-lactoalbumina bovina a pH 6,5 usando espectroscopia UV-VIS. Os resultados obtidos mostram que os polióis usados estabilizam a holo-α-lactoalbumina num grado significativamente menor ao reportado para outras proteínas. Se sugere que este menor efeito de estabilização ocorre devido a que esta proteína apresenta um estado desnaturalizado parcialmente desdobrado.

Caracterización de la formulación de -Macroagregados de albúmina / Characterization of - macroaggregates formulation

RESUMEN En este trabajo se realizó la caracterización del proceso de formación y marcaje de los macroagregados de albúmina (MAA) con . Con este fin se efectuaron estudios comparativos para establecer el papel de la desnaturalización en presencia del durante la formulación de -Sn-MAA, para ello se utilizaron tres formulaciones. Se realizó un análisis estructural cualitativo de la albúmina sérica humana para describir el sistema a través de un modelo gráfico. Los resultados mostraron diferencias importantes en el comportamiento radioquímico de las tres formulaciones. La formulación de -Sn-MAA (A) mostró un comportamiento superior, con purezas radioquímicas cercanas a 95% en los tiempos estudiados y una disociación inferior al 20% en 24 h.El modelo gráfico permitió explicar, a través de sencillas representaciones, los procesos involucrados en el sistema estudiado, permitiendo una mejor comprensión de este.

ABSTRACT The aim of this paper was to study the formation and radiolabelling of 99m Tcalbumin macroaggregates. Comparatives studies of three formulations to establish the role of denaturalization and in the formulation of -Sn-MAA were carried out. A qualitative structural analysis of human serum albumin was performed in order to give a description of the studied system using a graph model. The results showed important differences in the radiochemical behavior of the three formulations. -Sn-MAA (A) formulation had the best behavior, with radiochemical purity close to 95% and dissociation below 20% after 24 h of being radiolabeled. The graph model explained with simple representations the process involved in the studied system;thus allowing a better knowledge of it.

Structural similarities among the high molecular weight protein fractions of oilseeds.

Data on the physico-chemical properties of proteins from soybean, groundnut, sesame seed, sunflower seed, safflower seed, mustard seed, rapeseed and cotton seed are fairly extensive. An examination of the available data on high molecular weight proteins suggests that there are similarities in many of their properties. In this report the similarity in amino acid composition, size and shape, molecular weight, secondary structure, subunit composition, association-dissociation at high and low pH, stability towards denaturants, hydrolysis by enzymes and quaternary structure of the high molecular weight proteins is discussed. Based on these similarities a model has been proposed for the associationdissociation, denaturation and reassociation behaviour of the high molecular weight proteins of oilseeds.

Domain II + III of bovine serum albumin: Isolation and its characterization.

Some properties of a fragment of bovine serum albumin containing residues 184-582 of the protein sequence, produced by cyanogen bromide cleavage, have been reported. Urea-induced difference spectra of the fragment showed considerable exposure of aromatic chromophores by 8 Μ urea. Reversible unfolding of the fragment by urea, as followed by difference spectral measurements at 30°C, pH 7·0, occurred in two distinct steps involving at least 3 major conformational states, namely the native (N), intermediate (X) and completely denatured (D) states. The co-operativity values for the two transitions, N X and X Dwere found to be 4·0 and 16·4, respectively. Analysis of the data on bilirubin binding to bovine serum albumin and its fragment suggested that the fragment retains significant amount of its native structure. However, hydrodynamic parameters such as Stokes radius (3·14 nm), diffusion coefficient (6·98 × 10-7cm2/s) and frictional ratio (1·32) obtained by analytical gel chromatography as well as intrinsic viscosity (4·31 ml/g) indicates some asymmetry in the fragment molecule.