In Vitro Motility System to Study the Role of Motor Proteins in Receptor-Ligand Sorting

2007 ◽  
pp. 143-158 ◽  
Author(s):  
John W. Murray ◽  
Allan W. Wolkoff
Keyword(s):  
Motor Proteins ◽  
Receptor Ligand ◽  
In Vitro Motility

Translocation of microtubules caused by the alphabeta, beta and gamma outer arm dynein subparticles of Chlamydomonas

1998 ◽  
Vol 111 (9) ◽  
pp. 1155-1164 ◽  
Author(s):  
H. Sakakibara ◽  
H. Nakayama
Keyword(s):  
Glass Surface ◽  
Motor Proteins ◽  
Potassium Acetate ◽  
Dependent Manner ◽  
Acetate Concentration ◽  
Tubulin Dimer ◽  
Coated Glass ◽  
Heavy Chains ◽  
In Vitro Motility

Three kinds of subparticles of Chlamydomonas outer-arm dynein containing the alphabeta, beta and gamma heavy chains were isolated and assayed for their activities to translocate microtubules in vitro. All of them had activities to form bundles of microtubules in solution in an ATP-dependent manner and, when adsorbed on an appropriate glass surface, translocated microtubules. The alphabeta subparticle readily translocated microtubules on a silicone-coated glass surface with a velocity of 4.6 micron/second at 1 mM ATP. The beta subparticle translocated microtubules after it had been preincubated with tubulin dimer and when the Brownian movement of microtubules was suppressed by addition of methylcellulose. The velocity was on average 0.7 micron/second. The gamma subparticle translocated microtubules after being preincubated with tubulin dimer and adsorbed onto a silicone-coated glass surface. The velocity was about 3.8 micron/second. The tubulin dimer appeared to facilitate in vitro motility by blocking the ATP-insensitive binding of dynein subparticles to microtubule. The alphabeta, beta and gamma subparticles were thus found to have different properties as motor proteins. In addition, these subparticles showed different dependencies upon the potassium acetate concentration. Hence the outer-arm dynein of Chlamydomonas is a complex of motor proteins with different properties.

scholarly journals Rab8 Regulates the Actin-based Movement of Melanosomes

2005 ◽  
Vol 16 (4) ◽  
pp. 1640-1650 ◽  
Author(s):  
Marion L. Chabrillat ◽  
Claire Wilhelm ◽  
Christina Wasmeier ◽  
Elena V. Sviderskaya ◽  
Daniel Louvard ◽  
...  
Keyword(s):  
Rab Proteins ◽  
Rab Gtpases ◽  
Wild Type ◽  
In Vitro Motility Assay ◽  
Actin Bundles ◽  
In Vitro Motility ◽  
Rab Protein

Rab GTPases have been implicated in the regulation of specific microtubule- and actin-based motor proteins. We devised an in vitro motility assay reconstituting the movement of melanosomes on actin bundles in the presence of ATP to investigate the role of Rab proteins in the actin-dependent movement of melanosomes. Using this assay, we confirmed that Rab27 is required for the actin-dependent movement of melanosomes, and we showed that a second Rab protein, Rab8, also regulates this movement. Rab8 was partially associated with mature melanosomes. Expression of Rab8Q67L perturbed the cellular distribution and increased the frequency of microtubule-independent movement of melanosomes in vivo. Furthermore, anti-Rab8 antibodies decreased the number of melanosomes moving in vitro on actin bundles, whereas melanosomes isolated from cells expressing Rab8Q67L exhibited 70% more movements than wild-type melanosomes. Together, our observations suggest that Rab8 is involved in regulating the actin-dependent movement of melanosomes.

A step forward in the sigma enigma: a role for chirality in the sigma1 receptor–ligand interaction?

MedChemComm ◽  
2015 ◽  
Vol 6 (1) ◽  
pp. 138-146 ◽  
Author(s):  
Daniela Rossi ◽  
Annamaria Marra ◽  
Marta Rui ◽  
Erik Laurini ◽  
Maurizio Fermeglia ◽  
...  
Keyword(s):  
In Silico ◽  
Receptor Interaction ◽  
Ligand Interaction ◽  
Receptor Ligand ◽  
Sigma1 Receptor

To investigate the role of chirality in the ligand–σ1 receptor interaction, a series of enantiomeric arylalkylaminoalcohols and arylpyrrolidinols was evaluated by means of both in silico and in vitro studies.

scholarly journals A mutant heterodimeric myosin with one inactive head generates maximal displacement

2003 ◽  
Vol 162 (3) ◽  
pp. 481-488 ◽  
Author(s):  
Neil M. Kad ◽  
Arthur S. Rovner ◽  
Patricia M. Fagnant ◽  
Peteranne B. Joel ◽  
Guy G. Kennedy ◽  
...  
Keyword(s):  
Optical Trap ◽  
Actin Binding ◽  
Wild Type ◽  
Heavy Meromyosin ◽  
Maximum Displacement ◽  
Binding Partner ◽  
In Vitro Motility ◽  
Weak Binding

Each of the heads of the motor protein myosin II is capable of supporting motion. A previous report showed that double-headed myosin generates twice the displacement of single-headed myosin (Tyska, M.J., D.E. Dupuis, W.H. Guilford, J.B. Patlak, G.S. Waller, K.M. Trybus, D.M. Warshaw, and S. Lowey. 1999. Proc. Natl. Acad. Sci. USA. 96:4402–4407). To determine the role of the second head, we expressed a smooth muscle heterodimeric heavy meromyosin (HMM) with one wild-type head, and the other locked in a weak actin-binding state by introducing a point mutation in switch II (E470A). Homodimeric E470A HMM did not support in vitro motility, and only slowly hydrolyzed MgATP. Optical trap measurements revealed that the heterodimer generated unitary displacements of 10.4 nm, strikingly similar to wild-type HMM (10.2 nm) and approximately twice that of single-headed subfragment-1 (4.4 nm). These data show that a double-headed molecule can achieve a working stroke of ∼10 nm with only one active head and an inactive weak-binding partner. We propose that the second head optimizes the orientation and/or stabilizes the structure of the motion-generating head, thereby resulting in maximum displacement.

The role of silicon in forages: A quantitative X-Ray study

1987 ◽  
Vol 45 ◽  
pp. 416-417
Author(s):  
Janet H. Woodward ◽  
D. E. Akin
Keyword(s):  
Sodium Acetate ◽  
Dry Matter ◽  
Acetate Buffer ◽  
Plant Tissues ◽  
Sodium Acetate Buffer ◽  
X Ray ◽  
Dry Matter Digestibility ◽  
Percent Composition

Silicon (Si) is distributed throughout plant tissues, but its role in forages has not been clarified. Although Si has been suggested as an antiquality factor which limits the digestibility of structural carbohydrates, other research indicates that its presence in plants does not affect digestibility. We employed x-ray microanalysis to evaluate Si as an antiquality factor at specific sites of two cultivars of bermuda grass (Cynodon dactvlon (L.) Pers.). “Coastal” and “Tifton-78” were chosen for this study because previous work in our lab has shown that, although these two grasses are similar ultrastructurally, they differ in in vitro dry matter digestibility and in percent composition of Si.Two millimeter leaf sections of Tifton-7 8 (Tift-7 8) and Coastal (CBG) were incubated for 72 hr in 2.5% (w/v) cellulase in 0.05 M sodium acetate buffer, pH 5.0. For controls, sections were incubated in the sodium acetate buffer or were not treated.

An integrated screening method for evaluating the pulmonary toxicity of inhaled particulates: Role of microscopic techniques in assessing pulmonary macrophage clearance functions

1990 ◽  
Vol 48 (3) ◽  
pp. 826-827
Author(s):  
David B. Warheit ◽  
Lena Achinko ◽  
Mark A. Hartsky
Keyword(s):  
Bronchoalveolar Lavage ◽  
Pulmonary Toxicity ◽  
Screening Method ◽  
Pulmonary Response ◽  
Inhaled Particles ◽  
Cytochemical Analysis ◽  
Pulmonary Macrophages ◽  
Integrated Screening

There is a great need for the development of a rapid and reliable bioassay to evaluate the pulmonary toxicity of inhaled particles. A number of methods have been proposed, including lung clearance studies, bronchoalveolar lavage analysis, and in vitro cytotoxicity tests. These methods are often limited in scope inasmuch as they measure only one dimension of the pulmonary response to inhaled, instilled or incubated dusts. Accordingly, a comprehensive approach to lung toxicity studies has been developed.To validate the method, rats were exposed for 6 hours or 3 days to various concentrations of either aerosolized alpha quartz silica (Si) or carbonyl iron (CI) particles. Cells and fluids from groups of sham and dust-exposed animals were recovered by bronchoalveolar lavage (BAL). Alkaline phosphatase, LDH and protein values were measured in BAL fluids at several time points postexposure. Cells were counted and evaluated for viability, as well as differential and cytochemical analysis. In addition, pulmonary macrophages (PM) were cultured and studied for morphology, chemotaxis, and phagocytosis by scanning electron microscopy.

The Role of Polyphenols in the Modulation of Plasma Non-Enzymatic Antioxidant Capacity (NEAC)

2012 ◽  
Vol 82 (3) ◽  
pp. 228-232 ◽  
Author(s):  
Mauro Serafini ◽  
Giuseppa Morabito
Keyword(s):  
Antioxidant Capacity ◽  
Dietary Intervention ◽  
Ex Vivo ◽  
The Body ◽  
Dietary Polyphenols ◽  
Enzymatic Antioxidant ◽  
Endogenous Antioxidant

Dietary polyphenols have been shown to scavenge free radicals, modulating cellular redox transcription factors in different in vitro and ex vivo models. Dietary intervention studies have shown that consumption of plant foods modulates plasma Non-Enzymatic Antioxidant Capacity (NEAC), a biomarker of the endogenous antioxidant network, in human subjects. However, the identification of the molecules responsible for this effect are yet to be obtained and evidences of an antioxidant in vivo action of polyphenols are conflicting. There is a clear discrepancy between polyphenols (PP) concentration in body fluids and the extent of increase of plasma NEAC. The low degree of absorption and the extensive metabolism of PP within the body have raised questions about their contribution to the endogenous antioxidant network. This work will discuss the role of polyphenols from galenic preparation, food extracts, and selected dietary sources as modulators of plasma NEAC in humans.

Anti-obesity role of Aster glehni extract: in vivo and in vitro effects

Planta Medica ◽  
2012 ◽  
Vol 78 (11) ◽  
Author(s):  
HM Lee ◽  
TG Ahn ◽  
CW Kim ◽  
HJ An
Keyword(s):  
In Vitro Effects
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