Hydrolysis of fish oil by hyperactivated rhizomucor miehei lipase immobilized by multipoint anion exchange

Rhizomucor miehei lipase (RML) is greatly hyperactivated (around 20‐ to 25‐fold toward small substrates) in the presence of sucrose laurate. Hyperactivation appears to be an intramolecular process because it is very similar for soluble enzymes and covalently immobilized derivatives. The hyperactivated enzyme was immobilized (in the presence of sucrose laurate) on cyanogen bromide‐activated Sepharose (very mild covalent immobilization through the amino terminal residue), on glyoxyl Sepharose (intense multipoint covalent immobilization through the region with the highest amount of Lys residues), and on different anion exchangers (by multipoint anionic exchange through the region with the highest density of negative charges). Covalent immobilization does not promote the fixation of the hyperactivated enzyme, but immobilization on Sepharose Q retains the hyperactivated enzyme even in the absence of a detergent. The hydrolysis of fish oils by these hyperactivated enzyme derivatives was sevenfold faster than by covalently immobilized derivatives and three and a half times faster than by the enzyme hyperactivated on octyl‐Sepharose. The open structure of the hyperactivated lipase is fairly exposed to the medium, and no steric hindrance should interfere with the hydrolysis of large substrates. These new hyperactivated derivatives seem to be more suitable for hydrolysis of oils by RML immobilized inside porous supports. In addition, the hyperactivated derivatives are fairly stable against heat and organic cosolvents. © 2011 American Institute of Chemical Engineers Biotechnol. Prog., 2011     

The biological effects in animals related to the accident at the Chernobyl Atomic Electric Power Station. 1. The experimental model. The radiation loads on animals kept continuously under external and internal radiation exposure in the area of the Chernobyl Atomic Electric Power Station]

Irradiation conditions in which laboratory animals were kept in experimental laboratories of Chernobyl and Kiev after the accident at the Chernobyl A.P.S. are described. The data are presented on the spectral structural and activity of radionuclides in the diet as well as in the organs and tissues of the animals. The radiation loads have been estimated with regard to an external gamma component and the internal one contributed by the incorporated radionuclides. It has been shown that radiation doses received by the animals during their lifetime due to these contributions do not exceed units of cGy.     

The biological action of low-level radiation

On the basis of their own data and the literature the authors have analyzed the mechanism of biological action of low-level radiation on cells. In studying the harmful effect of gamma-radiation (10-40 cGy) on HeLa cells it was shown that damages occurred in 12 postirradiation generations. Analysis of cell distribution within the colonies has demonstrated that the share of colonies with low cell content and the number of giant cell increase. These data indicate that low-level radiation causes reproductive cell death that can occur not immediately but in later generations.     

Modelling of the disulphide-swapped isomer of human insulin-like growth factor-1: implications for receptor binding.

Insulin-like growth factor-1 (IGF-1) is a serum protein which unexpectedly folds to yield two stable tertiary structures with different disulphide connectivities; native IGF-1 [18-61,6-48,47-52] and IGF-1 swap [18-61,6-47, 48-52]. Here we demonstrate in detail the biological properties of recombinant human native IGF-1 and IGF-1 swap secreted from Saccharomyces cerevisiae. IGF-1 swap had a approximately 30 fold loss in affinity for the IGF-1 receptor overexpressed on BHK cells compared with native IGF-1.The parallel increase in dose required to induce negative cooperativity together with the parallel loss in mitogenicity in NIH 3T3 cells implies that disruption of the IGF-1 receptor binding interaction rather than restriction of a post-binding conformational change is responsible for the reduction in biological activity of IGF-1 swap. Interestingly, the affinity of IGF-1 swap for the insulin receptor was approximately 200 fold lower than that of native IGF-1 indicating that the binding surface complementary to the insulin receptor (or the ability to attain it) is disturbed to a greater extent than that to the IGF-1 receptor. A 1.0 ns high-temperature molecular dynamics study of the local energy landscape of IGF-1 swap resulted in uncoiling of the first A-region alpha-helix and a rearrangement in the relative orientation of the A- and B-regions. The model of IGF-1 swap is structurally homologous to the NMR structure of insulin swap and CD spectra consistent with the model are presented. However, in the model of IGF-1 swap the C-region has filled the space where the first A-region alpha-helix has uncoiled and this may be hindering interaction of Val44 with the second insulin receptor binding pocket.     

Effect of defrosting on quantity of volatile amines in stored sardines and herring used for tuna fattening

Studied were the quantities of volatile amines, total volatile bases nitrogen (TVB‐N) and trimethylamine nitrogen (TMA‐N) in stored sardines (Sardina pilchardus) and herring (Clupea harengus) over a 24‐h period after defrosting. At the beginning of the experiment analyses showed TVB‐N quantities in the flesh of both species to be higher than the FAO‐recommended limit. TMA‐N concentrations exceeded the acceptance limit 6 h after defrosting in herring samples and after 12 h in sardines. During the entire experiment TMA‐N and TVB‐N concentrations in sardines were above the tolerance limit. The defrosting method usually applied in tuna farms is unsuitable for maintenance of a high quality food supply. Low quality baitfish can cause reduced weight gain as well as induce health problems and mortality in farmed tuna (Thunnus thynnus) populations.     

Comparison of two techniques for quantifying environmental contaminants in human serum

Peak area matching and linear regression were used to quantify eight chlorinated pesticides and polychlorinated biphenyls (as Aroclor 1260) in human serum. There are no statistically significant differences in data obtained by these two quantifying techniques which were indicated by the paired t-test. For chlorinated pesticides, p = 0.053-0.62, and for polychlorinated biphenyls (as Aroclor 1260), p = 0.64. Analyte residues for the chlorinated pesticides ranged from 0.5 ppb for hexachlorobenzene (HCB) to 186 ppb for dichlorodiphenyldichloroethylene (DDE). Analyte residues for the polychlorinated biphenyls (as Aroclor 1260) ranged from 5-114 ppb. The absolute mean percent difference between the two quantifying techniques ranged from 0.06% for DDE to 8.06% for dieldrin (HEOD) among the chlorinated pesticides. The absolute mean percent difference between the two quantifying techniques for the polychlorinated biphenyls (as Aroclor 1260) was 3.4%. Peak area matching and linear regression were found to be comparable for quantifying these environmental residues in serum when the following conditions apply: 1) the concentration of the chlorinated pesticides is greater than or equal to 0.5 ppb (e.g., HCB, hexachlorocyclohexane (HCCH), oxychlordane (OC), heptachlor epoxide (HE), transnonachlor (TN), HEOD, and dichlorodiphenyltrichloroethane (DDT); 2) the concentration of the chlorinated pesticide is greater than or equal to 3 ppb (e.g., DDE); and 3) the total concentration of polychlorinated biphenyls (e.g., as Aroclor 1260) is greater than or equal to 5 ppb.     

In vivo protein synthesis from 14C-substrates in porcine tissues during the transition to postnatal development

[2-14C] leucine, [1-14C] alanine, [1-14C] glucose, [1-14C] lactate and [1-14C] pyruvate utilization in the protein synthesis has been studied in vivo at early stages of postnatal development of piglets. It has been established, that during the first 24 hours after birth the protein synthesis intensity, judging by [2-14C] leucine incorporation, in liver, skeletal muscle, duodenal wall and subcutaneous tissue of piglets increases 5, 7, 6.5 and 2.1 times respectively. At the age of 1-2 h the radioactive carbon incorporation from [1-14C] glucose into the brain proteins is more pronounced than into the proteins of liver and skeletal muscle. During the first days of life the intensity of the label incorporation from [1-14C] glucose into liver and skeletal muscle proteins of piglets is enhanced, whereas in brain it remains at the same level. The degree of 14C carbon incorporation from [1-14C]-alanine, [1-14C] pyruvate and [1-14C] lactate into the liver and skeletal muscle proteins of 5-days-old piglets is approximately the same, 14C substrates of protein synthesis in brain and subcutaneous adipose tissue having some peculiarities.     

Isolation of Actinomycetes synthesizing proteases with thrombolytic activity

Proteases with the thrombolytic activity were studied in 212 strains of actinomycetes isolated from different soils of the Soviet Union. The cultures belonged to the genera Micromonospora, Nocardia and Streptomyces. Proteases were synthesized by 41% of the studied actinomycetes and some of their strains completely dissolved in vitro artificially obtained blood thrombi within 120-240 min. In the Streptomyces genus, more active strains were found in the groups Flavus, Fradia and Globisporus. The groups Olivaceus, Violaceus and Viridis had less active strains.