Multiple Copies Of Protein Research Articles

Monosomes translate proteins in neurons

Neuroscience Like all other cells, neurons use different proteins to process information and respond to stimuli. To meet the huge demands for new proteins in their large and complex cell volume, neurons have moved the protein templates—messenger RNAs (mRNAs)—and the protein synthesis machines—ribosomes—out to synapses to make proteins locally. During protein synthesis, multiple ribosomes can form a structure known as a polysome, which produces multiple protein copies from a single mRNA. Working in rodent preparations, Biever et al. found that solitary, mRNA-associated ribosomes, or monosomes, are a substantial source of proteins in neuronal processes. Many synaptic proteins are made on single ribosomes, which may solve the problem of limited space in tiny synaptic compartments. Science , this issue p. [eaay4991][1] [1]: /lookup/doi/10.1126/science.aay4991

Old dogs with new tricks: intra-axonal translation of nuclear proteins.

Many different types of polarized eukaryotic cells have been shown to segregate synthesis for some protein subpopulations to cytoplasmic domains distant from their nucleus. For neurons, these distances can be tens-to-thousands fold more than the diameter of the cell body. Both axons and dendrites make use of this localized protein synthesis to bring autonomy to these far reaches of the cytoplasm (Gomes et al., 2014). This local mRNA translation is often used to mount a rapid response to extracellular stimuli encountered by the distal axon and dendrite. Indeed, activating translation of mRNAs residing locally at the synapse or growth cone brings a much more rapid response than could be achieved by transporting new proteins from the cell body. The neuron likely reaps a cost benefit from this mechanism in terms of energy consumption, since multiple protein copies can be generated from a single mRNA through sequential rounds of translation. Localized protein synthesis could also more effectively position a protein near its site of action or even bring an unanticipated novel function to the protein.

SNAP Dendrimers: Multivalent Protein Display on Dendrimer‐Like DNA for Directed Evolution

Display systems connect a protein with the DNA encoding it. Such systems (e.g., phage or ribosome display) have found widespread application in the directed evolution of protein binders and constitute a key element of the biotechnological toolkit. In this proof-of-concept study we describe the construction of a system that allows the display of multiple copies of a protein of interest in order to take advantage of avidity effects during affinity panning. To this end, dendrimer-like DNA is used as a scaffold with docking points that can join the coding DNA with multiple protein copies. Each DNA construct is compartmentalised in water-in-oil emulsion droplets. The corresponding protein is expressed, in vitro, inside the droplets as a SNAP-tag fusion. The covalent bond between DNA and the SNAP-tag is created by reaction with dendrimer-bound benzylguanine (BG). The ability to form dendrimer-like DNA straightforwardly from oligonucleotides bearing BG allowed the comparison of a series of templates differing in size, valency and position of BG. In model selections the most efficient constructs show recoveries of up to 0.86 % and up to 400-fold enrichments. The comparison of mono- and multivalent constructs suggests that the avidity effect enhances enrichment by up to fivefold and recovery by up to 25-fold. Our data establish a multivalent format for SNAP-display based on dendrimer-like DNA as the first in vitro display system with defined tailor-made valencies and explore a new application for DNA nanostructures. These data suggest that multivalent SNAP dendrimers have the potential to facilitate the selection of protein binders especially during early rounds of directed evolution, allowing a larger diversity of candidate binders to be recovered.