Video Experimental Relacionado
Updated: May 10, 2026
10:59
Artificial RNA Polymerase II Elongation Complexes for Dissecting Co-transcriptional RNA Processing Events
Published on: May 13, 2019
Cuantificación del ritmo de alargamiento durante la síntesis de proteínas de longitud completa.
Gabriel Rosenblum1, Chunlai Chen, Jaskiran Kaur
1Department of Chemistry, University of Pennsylvania, Philadelphia, Pennsylvania 19104-6323, USA.
Journal of the American Chemical Society
|July 5, 2013
Resumen
Las pausas ribosomales controlan el ritmo de síntesis de proteínas. La transferencia de energía de resonancia Förster de una sola molécula (smFRET) revela cómo la selección del ARN de transferencia (ARNt) influye en la velocidad y el ritmo de la síntesis de proteínas, lo que afecta el plegamiento y la modificación de las proteínas.
Área de la Ciencia:
- Biología Molecular Biología Molecular
- La biofísica es la biofísica.
- Genética La genética.
Sus antecedentes:
- Las pausas ribosómicas son críticas para regular la síntesis de proteínas y garantizar la formación adecuada de proteínas.
- Las mutaciones que afectan a estas pausas pueden conducir a proteínas mal plegadas y enfermedades humanas.
- Los métodos de conjunto tradicionales luchan por analizar pausas sutiles, lo que requiere técnicas avanzadas de una sola molécula.
Objetivo del estudio:
- Investigar el papel del uso del codón y el ARN de transferencia (ARNt) en la modulación de las tasas de alargamiento de la síntesis de proteínas.
- Evaluar cuantitativamente el impacto de la abundancia de tRNA en la dinámica de la síntesis de proteínas.
- Para explorar la influencia de la selección de tRNA en los procesos de co-traducción como el plegamiento y la modificación de proteínas.
Principales métodos:
- Utilizó la transferencia de energía de resonancia Förster de una sola molécula (smFRET) para monitorear la expresión de proteínas en tiempo real.
- Estudió la síntesis de una proteína de cuerpo entero, específicamente la variante de la proteína fluorescente verde esmeralda (GFP).
- Las tasas de alargamiento medidas se correlacionan con el uso de codones y la disponibilidad de ARNt de isoaceptor.
Principales resultados:
- Se demostraron correlaciones significativas entre las tasas de alargamiento de proteínas y el uso de codones específicos y sus tRNAs congénitos.
- Se estimó cuantitativamente el efecto de la sustitución de codones de reconocimiento de tRNA abundantes por codones de tRNA sinónimos y más raros en la tasa de alargamiento.
- Proporcionó evidencia para la selección de tRNA como un modulador clave de las tasas y ritmos de síntesis de proteínas.
Conclusiones:
- La selección de tRNA es un factor crucial en la regulación de la velocidad y el ritmo de la síntesis de proteínas.
- Estos hallazgos sugieren que la disponibilidad de tRNA puede influir en eventos simultáneos de co-traducción, incluido el plegamiento de proteínas y modificaciones químicas.
- Los enfoques de una sola molécula son efectivos para caracterizar pausas sutiles en la síntesis de proteínas que afectan la función celular y la enfermedad.
Videos de Conceptos Relacionados
02:35Transcription Elongation Factors
Transcription elongation is a dynamic process that alters depending upon the sequence heterogeneity of the DNA being transcribed. Hence, it is not surprising that the elongation complex's composition also varies along the way while transcribing a gene.
The transcription elongation is regulated via pausing of RNA polymerase on several occasions during transcription. In bacteria, these halts are necessary because the transcription of DNA into mRNA is coupled to the translation of that mRNA into a...
The transcription elongation is regulated via pausing of RNA polymerase on several occasions during transcription. In bacteria, these halts are necessary because the transcription of DNA into mRNA is coupled to the translation of that mRNA into a...
02:07Improving Translational Accuracy
Base complementarity between the three base pairs of mRNA codon and the tRNA anticodon is not a failsafe mechanism. Inaccuracies can range from a single mismatch to no correct base pairing at all. The free energy difference between the correct and nearly correct base pairs can be as small as 3 kcal/ mol. With complementarity being the only proofreading step, the estimated error frequency would be one wrong amino acid in every 100 amino acids incorporated. However, error frequencies observed in...
02:28Leaky Scanning
During most eukaryotic translation processes, the small 40S ribosome subunit scans an mRNA from its 5' end until it encounters the first start AUG codon. The large 60S ribosomal subunit then joins the smaller one to initiate protein synthesis. The location of the translation initiation is largely determined by the nucleotides near the start codon as there may be multiple translation initiation sites present on the mRNA. Marilyn Kozak discovered that the sequence RCCAUGG (where R stands for...
01:23Stringent Response in E. coli
Bacterial growth is closely tied to nutrient availability, with cells proliferating exponentially under favorable conditions and entering a stationary phase when resources become scarce. This transition is mediated by a regulatory mechanism known as the stringent response, which allows bacteria to adapt to nutrient deprivation by modulating gene expression and metabolic activity.During nutrient scarcity, intracellular amino acid levels decline. It results in the accumulation of uncharged tRNAs...
02:24Ribosome Profiling
Ribosome profiling or ribo-sequencing is a deep sequencing technique that produces a snapshot of active translation in a cell. It selectively sequences the mRNAs protected by ribosomes to get an insight into a cell’s translation landscape at any given point in time.
Applications of ribosome profiling
Ribosome profiling has many applications, including in vivo monitoring of translation inside a particular organ or tissue type and quantifying new protein synthesis levels.
The technique helps...
Applications of ribosome profiling
Ribosome profiling has many applications, including in vivo monitoring of translation inside a particular organ or tissue type and quantifying new protein synthesis levels.
The technique helps...
01:59Lagging Strand Synthesis
During replication, the complementary strands in double-stranded DNA are synthesized at different rates. Replication first begins on the leading strand. Replication starts later, occurs more slowly, and proceeds discontinuously on the lagging strand.
There are several major differences between synthesis of the leading strand and synthesis of the lagging strand. 1) Leading strand synthesis happens in the direction of replication fork opening, whereas lagging strand synthesis happens in the...
There are several major differences between synthesis of the leading strand and synthesis of the lagging strand. 1) Leading strand synthesis happens in the direction of replication fork opening, whereas lagging strand synthesis happens in the...