Application of recombinant enzymes in the production of human insulin

In the present study, the cleavage activities of both recombinant and bovine enzymes （trypsin and carboxy peptidase B） were determined using proinsulin fusion protein. The proteolysis products were analyzed with HPLC. We found that recombinant enzymes had stronger activities and produced fewer by-products. In addition, recombinant enzymes were more efficient in the production of human insulin than bovine enzymes.

pH-Dependence of Manganese (II) Oxidation Reaction by Novel Wild-Type and Mutants Recombinant Phlebia radiata Manganese Peroxidase 3 (rPr-MnP3) Enzymes

The goal of this study was to determine whether mutation of the Mn-binding site of wild-type recombinant Phlebia radiata manganese peroxidase 3 affected the pH-dependence kinetic parameters. pH range investigated was 2.5 – 12.0. The catalytic efficiency of the mutant enzymes at high and low pH in comparison to the wild-type was investigated using standard rPr-MnP3 protocol. Wild-type recombinant Phlebia radiata MnP3 enzyme showed optimal activity with Mn (II) as substrate at pH 5.0 and remained moderately active (approximately 40%) in the pH range of 6.0 - 9.0. The rPr-MnP3 mutants’ maximum activity ranged between 5.5 and 8.0. Wild-type and mutants rPr-MnP3 enzymes exhibited a similar pH profile with optimum pH of 3.0 for ABTS oxidation. Mutation has severely decreased the catalytic efficiency for Mn (II) oxidation at pH 5.0. The rPr-MnP3 enzymes showed enhanced affinity for Mn (II) at alkaline pH and a more alkaline range for catalysis than ever reported for any Manganese Peroxidase. This study reveals that at higher pH, rPr-MnP3 can function with alternative ligands in the Mn (II) site and does not have an absolutely obligate requirement for an all carboxylate ligand set. These results further strongly confirm that Mn2+ binding site is the only productive catalytic site for Mn (II) oxidation.

Termination of DNA Replication and the Role of Enzymes in Recombination

DNA is the genetic material of all cells, containing coded information about cellular molecules and processes. DNA consists of two polynucleofide strands twisted around each other in a double helix. The first step in cellular division is to replicate DNA so that copies can be distributed to daughter cells. Additionally, DNA is involved in transcribing proteins that direct cell growth and activities. However, DNA is tightly packed into genes and chromosomes. In order for replication or transcription to take place, DNA must firstly unpack itself so that it can interact with enzymes. DNA packing can be visualized as two very long strands that have been intertwined millions of times, tied into knots, and subjected to successive coiling. However, replication and transcription are much easier to accomplish if the DNA is neatly arranged rather than tangled up in knots. Enzymes are essential to unpacking DNA. Enzymes act to slice through individual knots and reconnect strands in a more orderly way. Hypothesizing that Termination of DNA replication proteins gave rise to those of eukaryotes during evolution, we chose the DNA polymerase （which infects microalgae） as the basis of this analysis, as it represents a primitive recombination. We show that it has significant similarity with replicative DNA polymerases of eukaryotes and certain of their large DNA. Sequence alignment confirms this similarity and establishes the presence of highly conserved domains in the polymerase amino terminus. Subsequent reconstruction of a phylogenetic tree indicates that these algal DNA are near the root of the containing all recombination. DNA polymerase delta members but that this does not contain the polymerases of other DNA. We consider arguments for the polarity of this relationship and present the hypothesis that the replication genes of DNA. DNA can be visualized as a complicated knot that must be unknotted by enzymes in order for replication or transcription to occur. It is perhaps not surprising then that connections between mathematical knot theory and biology have been discovered. By thinking of DNA as a knot, we can use knot theory to estimate how hard DNA is to unknot. This can help us estimate properties of the enzymes that unknot DNA.

Alternatives to antibiotics as growth promoters for use in swine production:a review

In the past two decades, an intensive amount of research has been focused on the development of alternatives to antibiotics to maintain swine health and performance. The most widely researched alternatives include probiotics, prebiotics, acidiflers, plant extracts and neutraceuticals such as copper and zinc. Since these additives have been more than adequately covered in previous reviews, the focus of this review will be on less traditional alternatives. The potential of antimicrobial peptides, clay minerals, egg yolk antibodies, essential oils, eucalyptus oil-medium chain fatty acids, rare earth elements and recombinant enzymes are discussed. Based on a thorough review of the literature, it is evident that a long and growing list of compounds exist which have been tested for their ability to replace antibiotics as feed additives in diets fed to swine. Unfortunately, the vast majority of these compounds produce inconsistent results and rarely equal antibiotics in their effectiveness. Therefore, it would appear that research is still needed in this area and that the perfect alternative to antibiotics does not yet exist.

The metabolism and hepatotoxicity of ginkgolic acid(17:1) in vitro

Pharmacological activities and adverse side effects of ginkgolic acids(GAs), major components in extracts from the leaves and seed coats of Ginkgo biloba L, have been intensively studied. However, there are few reports on their hepatotoxicity. In the present study, the metabolism and hepatotoxicity of GA(17:1), one of the most abundant components of GAs, were investigated. Kinetic analysis indicated that human and rat liver microsomes shared similar metabolic characteristics of GA(17:1) in phase I and II metabolisms. The drug-metabolizing enzymes involved in GA(17:1) metabolism were human CYP1 A2, CYP3 A4, UGT1 A6, UGT1 A9, and UGT2 B15, which were confirmed with an inhibition study of human liver microsomes and recombinant enzymes. The MTT assays indicated that the cytotoxicity of GA(17:1) in HepG2 cells occurred in a time-and dose-dependent manner. Further investigation showed that GA(17:1) had less cytotoxicity in primary rat hepatocytes than in HepG2 cells and that the toxicity was enhanced through CYP1 A-and CYP3 A-mediated metabolism.