scholarly journals Large-scale production and purification of the human green cone pigment: characterization of late photo-intermediates

1998 ◽  
Vol 330 (3) ◽  
pp. 1201-1208 ◽  
Author(s):  
Peter M. A. M. VISSERS ◽  
Petra H. M. BOVEE-GEURTS ◽  
M. Daniël PORTIER ◽  
Corné H. W. KLAASSEN ◽  
Willem J. DEGRIP
Keyword(s):  
Large Scale ◽  
Ph Dependence ◽  
Spectral Characteristics ◽  
Recombinant Baculovirus ◽  
Single Step ◽  
Scale Production ◽  
Spectral Position ◽  
Cone Pigment ◽  
Lipid Environment

We present the first characterization of the late photo-intermediates (Meta I, Meta II and Meta III) of a vertebrate cone pigment in a lipid environment. Marked differences from the same pathway in the rod pigment were observed. The histidine-tagged human green cone pigment was functionally expressed in large-scale suspension cultures in Sf9 insect cells using recombinant baculovirus. The recombinant pigment was extensively purified in a single step by immobilized metal affinity chromatography and displays the expected spectral characteristics. The purified pigment was able to activate the rod G-protein transducin at about half the rate of the rod pigment. Following reconstitution into bovine retina lipid proteoliposomes, identification and analysis of the photo-intermediates Meta I, Meta II and Meta III was accomplished. Similar to the rod pigment, our results indicate the existence of a Meta I-Meta II equilibrium, but we find no evidence for pH dependence. Replacement of native Cl- by NO3- in the anion-binding site of the cone pigment affected the spectral position of the pigment itself and of the Meta I intermediate, but not that of Meta II and Meta III. The decay rate of the ‘active’ intermediate Meta II did not differ for the Cl- and NO3- state. However, in qualitative agreement with results reported before for chicken cone pigments, the rate of Meta II decay was significantly higher in the human cone pigment than in the rod pigment.

scholarly journals Large-scale production and purification of functional recombinant bovine rhodopsin with the use of the baculovirus expression system

1999 ◽  
Vol 342 (2) ◽  
pp. 293-300 ◽  
Author(s):  
Corné H. W. KLAASSEN ◽  
Petra H. M. BOVEE-GEURTS ◽  
Godelieve L. J. DECALUWÉ ◽  
Willem J. DEGRIP
Keyword(s):  
Membrane Proteins ◽  
Large Scale ◽  
Structural Changes ◽  
Expression System ◽  
Recombinant Baculovirus ◽  
Baculovirus Expression System ◽  
Scale Production ◽  
Protein Activation ◽  
Bovine Rhodopsin ◽  
Lipid Environment

Here we describe a generic procedure for the expression and purification of milligram quantities of functional recombinant eukaryotic integral membrane proteins, exemplified by hexahistidine-tagged bovine rhodopsin. These quantities were obtained with the recombinant baculovirus/Sf9 insect cell-based expression system in large-scale bioreactor cultures with the use of a serum-free and protein-free growth medium. After optimization procedures, expression levels up to 4 mg/l were established. The recombinant rhodopsin could be purified with high overall yield by using immobilized-metal-affinity chromatography on Ni2+-agarose. After reconstitution into a native lipid environment, the purified protein was functionally indistinguishable from native rhodopsin with regard to the following parameters: spectral absorbance band, structural changes after photoactivation, and G-protein activation. The procedures developed can be adapted to other membrane proteins. The ability to produce and purify tens of milligrams of functional recombinant eukaryotic membrane protein meets the ever-increasing demand of material necessary to perform detailed biochemical and structural biophysical studies that are essential in unravelling their working mechanism at a molecular level.

scholarly journals Large-scale production and physicochemical characterization of human immune interferon.

1979 ◽  
Vol 26 (1) ◽  
pp. 36-41 ◽  
Author(s):  
M P Langford ◽  
J A Georgiades ◽  
G J Stanton ◽  
F Dianzani ◽  
H M Johnson
Keyword(s):  
Large Scale ◽  
Physicochemical Characterization ◽  
Scale Production ◽  
Immune Interferon ◽  
Large Scale Production ◽  
Human Immune

scholarly journals Electrochemically derived functionalized graphene for bulk production of hydrogen peroxide

2020 ◽  
Vol 11 ◽  
pp. 432-442 ◽  
Author(s):  
Munaiah Yeddala ◽  
Pallavi Thakur ◽  
Anugraha A ◽  
Tharangattu N Narayanan
Keyword(s):  
Large Scale ◽  
Reduction Reaction ◽  
Single Step ◽  
Scale Production ◽  
Functionalized Graphene ◽  
Large Scale Production ◽  
Production Of Hydrogen ◽  
Electron Reduction ◽  
Bulk Production ◽  
Carbon Based

On-site peroxide generation via electrochemical reduction is gaining tremendous attention due to its importance in many fields, including water treatment technologies. Oxidized graphitic carbon-based materials have been recently proposed as an alternative to metal-based catalysts in the electrochemical oxygen reduction reaction (ORR), and in this work we unravel the role of C=O groups in graphene towards sustainable peroxide formation. We demonstrate a versatile single-step electrochemical exfoliation of graphite to graphene with a controllable degree of oxygen functionalities and thickness, leading to the formation of large quantities of functionalized graphene with tunable rate parameters, such as the rate constant and exchange current density. Higher oxygen-containing exfoliated graphene is known to undergo a two-electron reduction path in ORR having an efficiency of about 80 ± 2% even at high overpotential. Bulk production of H2O2 via electrolysis was also demonstrated at low potential (0.358 mV vs RHE), yielding ≈34 mg/L peroxide with highly functionalized (≈23 atom %) graphene and ≈16 g/L with low functionalized (≈13 atom %) graphene, which is on par with the peroxide production using state-of-the-art precious-metal-based catalysts. Hence this method opens a new scheme for the single-step large-scale production of functionalized carbon-based catalysts (yield ≈45% by weight) that have varying functionalities and can deliver peroxide via the electrochemical ORR process.

scholarly journals Preclinical Characterization of Naturally Occurring Polyketide Cyclophilin Inhibitors from the Sanglifehrin Family

2011 ◽  
Vol 55 (5) ◽  
pp. 1975-1981 ◽  
Author(s):  
Matthew A. Gregory ◽  
Michael Bobardt ◽  
Susan Obeid ◽  
Udayan Chatterji ◽  
Nigel J. Coates ◽  
...  
Keyword(s):  
Large Scale ◽  
Scale Production ◽  
Subgenomic Replicon ◽  
Isomerase Activity ◽  
Lead Compounds ◽  
Naturally Occurring ◽  
Large Scale Production ◽  
Pharmacokinetic Properties ◽  
Biosynthetic Engineering

ABSTRACTCyclophilin inhibitors currently in clinical trials for hepatitis C virus (HCV) are all analogues of cyclosporine (CsA). Sanglifehrins are a group of naturally occurring cyclophilin binding polyketides that are structurally distinct from the cyclosporines and are produced by a microorganism amenable to biosynthetic engineering for lead optimization and large-scale production by fermentation. Preclinical characterization of the potential utility of this class of compounds for the treatment of HCV revealed that the natural sanglifehrins A to D are all more potent than CsA at disrupting formation of the NS5A-CypA, -CypB, and -CypD complexes and at inhibition of CypA, CypB, and CypD isomerase activity. In particular, sanglifehrin B (SfB) was 30- to 50-fold more potent at inhibiting the isomerase activity of all Cyps tested than CsA and was also shown to be a more potent inhibitor of the 1b subgenomic replicon (50% effective concentrations [EC50s] of 0.070 μM and 0.16 μM in Huh 5-2 and Huh 9-13 cells, respectively). Physicochemical and mouse pharmacokinetic analyses revealed low oral bioavailability (F< 4%) and low solubility (<25 μM), although the half-lives (t1/2) of SfA and SfB in mouse blood after intravenous (i.v.) dosing were long (t1/2> 5 h). These data demonstrate that naturally occurring sanglifehrins are suitable lead compounds for the development of novel analogues that are less immunosuppressive and that have improved metabolism and pharmacokinetic properties.

scholarly journals Characterization of graphene nanosheets obtained by a modified Hummer's method

2018 ◽  
Vol 23 (1) ◽  
Author(s):  
Renata Hack ◽  
Cláudia Hack Gumz Correia ◽  
Ricardo Antônio de Simone Zanon ◽  
Sérgio Henrique Pezzin
Keyword(s):  
Electron Microscopy ◽  
Large Scale ◽  
Graphene Nanosheets ◽  
Scale Production ◽  
Natural Graphite ◽  
Graphite Oxidation ◽  
Transmission Electron ◽  
Large Scale Production ◽  
High Degree

ABSTRACT Natural graphite is an inexpensive and abundant source to obtain graphene nanosheets. The most efficient method for large-scale production is the chemical method, which is based on the oxidation of natural graphite. This paper reports the synthesis and characterization of graphene obtained by the Hummers method with some modifications. The results indicate a high degree of graphite oxidation, proving that the process was efficient. Analyses of field emission scanning electron microscopy (FEG), transmission electron microscopy (TEM), Raman spectroscopy, thermogravimetric analysis (TGA) and X-ray diffraction showed that the graphene produced presented characteristics similar to the commercial graphene.

Growth and characterization of high-efficiency InGaN MQW blue and green LEDs from large-scale-production MOCVD reactors

1999 ◽  
Author(s):  
Chuong A. Tran ◽  
Robert F. Karlicek, Jr. ◽  
Michael G. Brown ◽  
Ivan Eliashevich ◽  
Alexander Gurary ◽  
...  
Keyword(s):  
Large Scale ◽  
High Efficiency ◽  
Scale Production ◽  
Large Scale Production ◽  
Green Leds
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