Ash Kothare
Professor Hanke
Honors Biology
19 September 2015
Paper 5
The paper, Function of Aggregated Reticulocyte Ribosomes in Protein Synthesis by A. Gierer, details the findings about a study to answer the question whether multiple ribosomes can operate on one molecule of messenger RNA simultaneously. Firstly, background information is necessary for understanding this paper. Ribosomes, the organelle where protein synthesis occurs, require a messenger RNA to be attached to it to be labelled active. Active ribosomes have an increased molecular weight due to the mRNA which in turn causes a higher sedimentation coefficient. The value of the sedimentation coefficient is calculated by timing the movement of a particle in a medium of known viscosity.
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Ribosomes are often labeled by their sedimentation coefficient. In this experiment, rabbit reticulocytes were used with a sedimentation coefficient of 80 s. The study focuses on the rapidly sedimenting fractions. These fast fractions are aggregates of ribosomes. According to this study most of the protein synthesized is of one type: hemoglobin. There are two hypotheses for this experiment. The first model is where mRNA is statically linked to the ribosomal surface. One ribosome operates on one molecule of mRNA. The ribosome would contain sites where a peptide bond could be formed because of the amino acids in the peptide chain. The growing peptide needs to be attached to the ribosome only at the point of growth which is why several peptides would not grow simultaneously on the same ribosome. The second model is where mRNA is not statically linked to the ribosome but is attached by a small group of nucleotides to the ribosomal surface. So there is one site of synthesis on the ribosome to which the peptide chain can be attached to at its point of growth. In this model, only one peptide can grow on each ribosome but several ribosomes can operate simultaneously on one molecule of messenger RNA. The control groups for this experiment are unfractionated ribosomes and the mixture without ribosomes. While the four experimental groups are [14C]valine without poly-U, [14C]valine with poly-U, [14C]phenylalanine without poly-U, and [14C]phenylalanine with poly-U. Here the poly-U acts as a mRNA. These four experimental groups will be compared to the 2 controls to determine the differences of fractionation in cts/min. An important technique used throughout this experiment is centrifuging.
A couple other techniques used were incubation, titration, sucrose gradient and measurement of radioactivity. Incubation was necessary to add the RNase to the cell so it would disintegrate into ribosomal units. Titration is the addition of one solution of a known molarity to a known volume of another solution of unknown concentration until the reaction reaches neutralization. Here the supernatant solution was titrated to a pH of 7.5. The purpose behind centrifuging the cell is to isolate the ribosomal particles from the cell lysates. The ribosomes are named by the sedimentation characteristics during centrifugation. The sedimentation properties of a particle depend on its molecular size and geometrical shape. Sucrose gradient centrifugation also known as isopycnic centrifugation, particles of a specific density sediment until they reach the point where their density is the same as the gradient media in this case the sucrose. The particles are then separated according to their …show more content…
buoyancy. According to the experiment the data showed that Model 2 was more likely. They concluded one mRNA molecule is made up of several sections and each code for a different peptide chain. The results indicate that many of the aggregated ribosomes contain mRNA, but are not receptive to added mRNA. On the other hand, unaggregated ribosomes do not contain mRNA, but are receptive to added messenger RNA molecules. The linkage of the ribosomes as aggregates is because of the messenger RNA. The findings support a theory of protein synthesis where multiple ribosomes operate simultaneously on one molecule of messenger RNA. There might be a correlation of the activity of higher aggregates in protein synthesis of reticulocytes with the fact that the main product of protein synthesis, the hemoglobin molecule, contains four peptide chains per molecule. The paper assumes that four ribosomes have to be assembled to produce and to release a hemoglobin molecule. This study brings up the questions of whether this correlation between peptide chains per molecule can be applied to other cells besides reticulocytes for protein synthesis of proteins besides hemoglobin, and what role could mRNA and ribosomes play in medicine. Presently there have been developments in this theory. An article written in 2008 discusses, “how my laboratory found that multiple ribosomes traverse each mRNA, yielding the structures known as polysomes”. A book written in 1997 summarizes that the current understanding of ribosomes and mRNA is that, “Translation of an mRNA molecule occurs in a multimeric structure, the polysome, which consists of multiple ribosomes arrayed along the length of the mRNA”. So Gierer’s paper concluding that protein synthesis occurs due to several ribosomes operating simultaneously on one molecule of messenger RNA was correct. Also there is a name given to what this is called: A polyribosome (or polysome) which is a complex of a mRNA molecule and two or more ribosomes that is formed during active translation. This information is used today for biochemical analysis of ribosome and polysome associated factors such as signaling molecules. Works Cited PROFILES, I.
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Living organisms undergo chemical reactions with the help of unique proteins known as enzymes. Enzymes significantly assist in these processes by accelerating the rate of reaction in order to maintain life in the organism. Without enzymes, an organism would not be able to survive as long, because its chemical reactions would be too slow to prolong life. The properties and functions of enzymes during chemical reactions can help analyze the activity of the specific enzyme catalase, which can be found in bovine liver and yeast. Our hypothesis regarding enzyme activity is that the aspects of biology and environmental factors contribute to the different enzyme activities between bovine liver and yeast.
Miller, Kenneth R. and Joseph S. Levine. “Chapter 12: DNA and RNA.” Biology. Upper Saddle River: Pearson Education, Inc., 2002. Print.
According to the graph on amylase activity at various enzyme concentration (graph 1), the increase of enzyme dilution results in a slower decrease of amylose percentage. Looking at the graph, the amylose percentage decreases at a fast rate with the undiluted enzyme. However, the enzyme dilution with a concentration of 1:3 decreased at a slow rate over time. Additionally, the higher the enzyme dilution, the higher the amylose percentage. For example, in the graph it can be seen that the enzyme dilution with a 1:9 concentration increased over time. However, there is a drastic increase after four minutes, but this is most likely a result of the error that was encountered during the experiment. The undiluted enzyme and the enzyme dilution had a low amylose percentage because there was high enzyme activity. Also, there was an increase in amylose percentage with the enzyme dilution with a 1: 9 concentrations because there was low enzyme activity.
Small ribosomes scattered throughout the cytoplasm. No mitochondria hence respiration takes place on an infolding of the cell membrane
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In our Biology Lab we did a laboratory experiment on fermentation, alcohol fermentation to be exact. Alcohol fermentation is a type of fermentation that produces the alcohol ethanol and CO2. In the experiment we estimated the rate of alcohol fermentation by measuring the rate of CO2 production. Both glycolysis and fermentation consist of a series of chemical reactions, each of which is catalyzed by a specific enzyme. Two of the tables substituted some of the solution glucose for two different types of solutions. They are as followed, Table #5 substituted glucose for sucrose and Table #6 substituted the glucose for pH4. The equation for alcohol fermentation consists of 6 Carbons 12 Hydrogens 6 Oxygen to produce 2 pyruvates plus 2 ATP then finally the final reaction will be 2 CO2 plus Ethanol. In the class our controlled numbers were at Table #1; their table had 15 mL Glucose, 10 mL RO water, and 10 mL of yeast which then they placed in an incubator at 37 degrees Celsius. We each then measured our own table’s fermentation flasks every 15 mins for an hour to compare to Table #1’s controlled numbers. At
mRNA is stabilized in the cytoplasm by adding a 5’ cap and a 3’ poly adenine tail, which prevents degradation by ribonucleases. The binding of miRNA can cause 3 events to occur; deadenylation, decapping, and 5’ to 3’ degradation1. Often in the 3’ untranslated region (UTR) there are AU rich elements (AREs) when the miRNA along with Argonaute 1, Argonaute 2 and Dicer1 is bound it allows rapid decay of the mRNA
Cells are a very important part of our bodies that are necessary for our survival. Within the cell there are many different organelles that have many different functions. One of those organelles are ribosomes, and they are responsible for causing Alopecia. Cells are filled with thousands of these tiny ribosomes that are responsible for producing protein molecules, which are needed for all life processes. (Storad, 1998 ). In the ribosomes, a codon from the mRNA is connected to the anticodon of the tRNA, and this starts forming amino acid chains, to make proteins. (Renneboog, 2014).
Some ribosomes are found scattered in the cytoplasm (these can be referred to as free ribosomes), while others are attached to the endoplasmic reticulum (also known as bound ribosomes). The surface of the endoplasmic reticulum when bound with ribosomes is called rough endoplasmic reticulum (RER). Both the free and bound ribosomes have similar structure and are responsible for the production of proteins. Ribosomes are responsible for assembling amino acids to form specific proteins, which in turn are essential for carrying out the cell's
There are huge numbers of genes in our genome yet only few of them express to synthesis mRNAs which encode different proteins. These mRNAs are collectively called as transcriptome and mRNA can be reverse transcribed into cDNA, which provides evidence for all mRNA transcripts. Hence, mRNA and cDNA are crucial for gene expression profiling and transcriptome study.
In 1998, the concept of RNA interference (RNAi) was first discovered and added to the complexity of post-transcriptional regulation of gene expression in cells (Fire, 1998). The RNAi phenomenon was originally discovered in Caenorhabditis elegans where the injection of double-stranded RNA resulted in the decreased expression of genes with highly homologous sequences to the injected nucleic acid sequence. In the first step of the mechanism of RNAi, double stranded RNA is converted cleaved into short, 21 to 24 nucleotide long small interfering RNAs (siRNAs) (Elbashir, 2001). RNA cleavage is catalyzed by the enzyme Dicer, an endonuclease of the RNase III family (Layzer, 2004). The resultant siRNAs contain 3'-hydroxyl termini and a 5'-phosphate at both ends. In the second step of RNAi, these siRNAs are incorporated into the RNA-induced silencing complex (RISC). Within the complex, a helicase unwinds the duplex siRNA and the resulting single-stranded siRNAs can pair with messenger RNAs (mRNAs) that contain a high degree of sequence complementarity to the siRNA. Following this in humans, the Argonaute 2 (Ago2) protein that is associated with the RISC complex degrades the targeted mRNA. The target mRNA is cleaved in the complementary region at the phosphodiester bond that lies across nucleotides 10 and 11 of the 5'-end of the siRNA (Elbashir, 2001). For RNAi-mediated cleavage and degradation of mRNA to be successful, a 5'-phosphate must be present on the antisense strand and the antisense-mRNA helical duplex must be in the A-form (Chiu, 2003.)
1.1 Non-coding RNAs The central dogma of molecular biology states that genetic information is conveyed from DNA to mRNA to protein implying that proteins are the main functional genetic output (Crick 1970). Even those few early known non-protein-coding RNAs (ncRNAs) such as transfer RNA, ribosomal RNA, snoRNAs and splicosomal RNAs were in the end required for mRNA processing and translation. The dogma might still be applicable to prokaryotes whose genome consists of approx. 90 % protein-coding genes. In eukaryotes, however, only about 2 % of the genes are protein-coding (Alexander et al. 2010) and those have been studied intensively. The remaining major fraction of the genomic output has for a long time been classified as genetic junk, as most transcripts had low or no protein-coding capacity nor cis-regulatory functions. Techniques like high resolution microarray and improved sequencing assays revealed that 98 % of the human genome consists of non-protein coding sequences compared to 25 % in prokaryotes. Remarkably, this increased proportion of ncRNAs (and not the number of protein-coding genes) comes along with higher developmental complexity. When proteins reach their functional limits, other regulatory components such as introns and other sequences coding for ncRNAs evolved (Mattick 2004). Coincident with the abundance of ncRNAs, higher species possess also more proteins carrying RNA- binding sites (Mattick & Makunin 2006). The demotion of non-coding transcripts as „transcriptional noise“ had to be corrected as a significant number of non-coding transcripts showed cell type-specific expression, specific localization in cellular compartments, functional relevance for development and p...
Campbell, N. A. & J. B. Reece, 8th eds. (2008). Biology. San Francisco: Pearson Benjamin Cummings.