Lipid Droplets Purified from Drosophila Embryos as an Endogenous Handle for Precise Motor Transport Measurements
| Autor: | George T. Shubeita, Rafael A. Longoria, Ernst-Ludwig Florin, Tobias F. Bartsch |
|---|---|
| Rok vydání: | 2013 |
| Předmět: |
Embryo
Nonmammalian Optical Tweezers Biophysics Analytical chemistry Motility Endogeny 02 engineering and technology Biology Microtubules 03 medical and health sciences Microtubule Lipid droplet Organelle Animals Molecular Machines Motors and Nanoscale Biophysics 030304 developmental biology 0303 health sciences Molecular Motor Proteins Vesicle Biological Transport Lipid Metabolism 021001 nanoscience & nanotechnology biology.organism_classification Drosophila melanogaster Optical tweezers 0210 nano-technology |
| Zdroj: | Biophysical Journal. 105:1182-1191 |
| ISSN: | 0006-3495 |
| Popis: | Molecular motor proteins are responsible for long-range transport of vesicles and organelles. Recent works have elucidated the richness of the transport complex, with multiple teams of similar and dissimilar motors and their cofactors attached to individual cargoes. The interaction among these different proteins, and with the microtubules along which they translocate, results in the intricate patterns of cargo transport observed in cells. High-precision and high-bandwidth measurements are required to capture the dynamics of these interactions, yet the crowdedness in the cell necessitates performing such measurements in vitro. Here, we show that endogenous cargoes, lipid droplets purified from Drosophila embryos, can be used to perform high-precision and high-bandwidth optical trapping experiments to study motor regulation in vitro. Purified droplets have constituents of the endogenous transport complex attached to them and exhibit long-range motility. A novel method to determine the quality of the droplets for high-resolution measurements in an optical trap showed that they compare well with plastic beads in terms of roundness, homogeneity, position sensitivity, and trapping stiffness. Using high-resolution and high-bandwidth position measurements, we demonstrate that we can follow the series of binding and unbinding events that lead to the onset of active transport. |
| Databáze: | OpenAIRE |
| Externí odkaz: |
|