Ribosomes are protein factories found in cells that translate mRNA into polypeptide chains. The first high-resolution crystal structures of ribosomes were published in the early 2000s by Ramakrishnan's team. These structures showed that ribosomes consist of two subunits that fit together during protein synthesis. Ramakrishnan's work helped reveal insights into ribosome evolution and translation mechanisms like how silent mutations in tRNA can affect decoding. Current projects aim to crystallize ribosomes in different functional states to better understand translation.

1. Ribosomes: Unravelling the Crystal structure of the
Protein Factories
Name: D.Sairam
Course: Bioorganic and Bioinorganic
Chemistry
Course Code: BSBT-211
Presentation Code: U1P1
Course Instructor : Dr. Vineet Sharma

2. Overview
• Introduction
• Evolutionary Description
• Crystal structure
• Translation in Ribosomes
• Mutations in tRNA
• Current Projects by Venkatraman Ramakrishnan
• References

3. Introduction
• Ribosomes are a prominent part of the cell. They are present in
both Eukaryotic and Prokaryotic cells.
• Some ribosomes attach to the Endoplasmic Reticulum and
some float freely in the cytoplasm.
• Whether Ribosomes are free or attached, they usually cluster in
groups connected by a strand of another kind of ribonucleic acid
called messenger RNA (mRNA).
• These clusters are called polyribosomes or polysomes.
• One can classify them into two major zones (On the basis of
function performed): First the region which reads and binds to
the RNA sequence and the Second part joins the amino acids to
form peptides.

4. • Ribosomes consist of two subunits that fit together and work
as one to translate the mRNA into a polypeptide chain during
protein synthesis.
• It is because they are formed from two subunits of non-equal
size, they are slightly longer in the axis than in diameter.
• Prokaryotic ribosomes are around 20 Nm in diameter and are
composed of 65% rRNA and 35% Ribosomal Proteins.
• Bacterial Ribosomes are composed of one or two rRNA
strands. Eukaryotic ribosomes contain one or three very large
rRNA molecules and multiple smaller protein. The ribosomal
subunits of Prokaryotes and Eukaryotes are quite similar.
• The unit of measurement is the Svedberg unit, a measure of
the rate of sedimentation in centrifugation rather than size.

5. Source: http://www.osti.gov/accomplishments/ramakrishnan_steitz.html

6. Evolutionary Description[1]
• The origins and evolution of the ribosome dates back to about 3–4
billion years ago and form the basis for biochemistry and the structure
of the ribosome.
• Processes of ribosomal RNA (rRNA) expansion can be “observed” by
comparing 3D rRNA structures of bacteria (small), yeast (medium),
and metazoans (large).
• rRNA’s size varies with species’ size and complexity.
• Differences in ribosomes across species reveal that rRNA expansion
segments have been added to rRNAs without disturbing the pre-
existing core.
• This study helps us provide insights into the ancestral structure of the
Cell. [2]

7. Crystal Structure[3]
• The general molecular structure of the ribosome has been known since
the early 1970s. In the early 2000s the structure has been achieved at
high resolutions, on the order of a few Angstrom. [4]
• The first papers giving the structure of the Ribosome at atomic
resolution were published almost simultaneously in early 2000s.
• The 50S (large prokaryotic) subunit was determined from the
Bacterium Deinococcus radiodurans and the structure of the 30S
subunit was determined from Thermus thermophiles.
• After two years of extensive research the crystal structure of the hybrid
state of Ribosomes was determined by Ramakrishnan and his team.
• A protein release factor (RF3) like Guanosine triphosphatase [GTP]
binds to the ribosome and promotes dissociation of release factors
during Peptide Synthesis.
• These structural studies were awarded the Nobel Prize in Chemistry in
2009.

8. Source:
http://www.pnas.org/content/108/38/15676.ful
l

9. Translation in Ribosomes
• Translation is a Biological process in which ribosomes make
proteins. In short there are four phases of Translation.
• Initiation: The ribosome assembles around the target mRNA.
This gets attached to a start codon such as Methionine.
• Elongation: The tRNA gets transferred to an amino acid and
subsequently to the corresponding codon.
• Translocation : The ribosome then moves or (Trans locates) to
the next mRNA codon to continue the process, creating an amino
acid chain.
• Termination: It occurs when a stop codon is reached, the
ribosome releases the polypeptide.
• Ramakrishnan’s team worked on initiation phase of Translation
in a Virus called Cricket Paralysis Virus or CPVs.

10. Translation in Cricket Paralysis Virus [5]
• The Cricket Paralysis Virus internal ribosome entry site
(CrPV-IRES) is a folded structure in a viral mRNA that allows
initiation of translation in the absence of any host initiation
factors.
• By using recent advances in single-particle Electron
Cryomicroscopy, they have solved the structure of CrPV-IRES
bound to the ribosome of the yeast (host) Kluyveromyces
lactis.
• The structure and accompanying factor-binding data show that
CrPV-IRES binding mimics a pre-Translocation phase rather
than initiation state of the ribosome.
• Translocation of the IRES by elongation factor 2 (eEF2) is
required to bring the first codon of the mRNA into the site and
to allow the start of translation.

11. Source: http://www.theguardian.com/science/2013/nov/24/venkatraman-ramakrishnan-
ribosome-nobel-chemistry

12. Mutations in tRNA
• Silent mutations are DNA mutations that do not significantly
alter the phenotype of the organism in which they occur. Silent
mutations may occur in non- coding regions (outside
of genes within Introns), or they may occur within Exons.
• When they occur within exons they either do not result in a
change to the Amino acid sequence of a protein (i.e.
Asynonymous substitution), or result in the insertion of an
alternative amino acid with similar properties to that of the
original amino acid.
• Silent mutations do not alter protein function they are often
treated as though they are evolutionary useful

13. • Transfer RNA (tRNA) availability is one of the reasons that silent
mutations might not be as silent as they are conventionally believed to
be.
• Ramakrishnan’s Team worked on how these silent mutations affect the
decoding factors while translations[6]

14. Current Research Projects by Team Ramakrishnan
• Ramakrishnan and his team are trying to crystallize the
ribosomes in complex with mRNA, tRNA and various factors in
functional states during initiation, elongation and termination.
• Another ongoing project is trying to understand the interactions
of initiation factors with the 40S ribosomal subunit, mRNA and
initiator tRNA, as well as the conformational changes induced by
the binding of these factors.

15. References
• http://www.ncbi.nlm.nih.gov/pubmed/19955257[1]
• http://www.pnas.org/content/111/28/10251.abstract[2]
• http://www.ncbi.nlm.nih.gov/pubmed/19955257[3]
• http://www.pnas.org/content/108/38/15798.abstract[4]
• http://dx.doi.org/10.1126%2Fscience.1060089[5]
• http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4017093[6]
• http://www.ncbi.nlm.nih.gov/pubmed/?term=How+mutations+in+t
RNA+distant+from+the+anticodon+affect+the+fidelity+of+decodi
ng
• http://www.weizmann.ac.il/sb/faculty_pages/Yonath/00Sc_activiti
es.html
• http://www.yale.edu/steitz/research.html
• http://www.ncbi.nlm.nih.gov/pubmed/16959973
• http://www.uniprot.org/uniprot/P60228

16. Thank You