Pamela Osborn Overview What are ribozymes Background Structure Hairpin Hammerhead and RNase P Future applications Function amp Role of ribozymes within the body Mechanism Role of metals in catalysis amp ID: 688050
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Slide1
Ribozymes: RNA Enzymes - Function & Role
Pamela OsbornSlide2
Overview
What
are ribozymes?
Background
Structure
Hairpin, Hammerhead, and
RNase
P
Future applications
Function
& Role of ribozymes within the body
Mechanism: Role of metals in catalysis &
nucleobase
acid/base catalysis
.
Kinetics: Efficiency of ribozymesSlide3
What Are Ribozymes?
Ribonucleic acid capable of catalyzing a chemical reaction.
Previously thought to be the work of proteins.
Vital for many biological processes
Natural ribozymes catalyze phosphodiester transfer or hydrolysis.
Thought to be remnants of an RNA dominated world.
Could solve the chicken or egg dilemmaSlide4
1989 Nobel Prize in Chemistry:
Sidney Altman & Thomas R.
Cech
They
showed that RNA was capable of
splicing
itself and joining the genetically important pieces together. By observing this behavior in unprocessed RNA molecules not in the presence of proteins, they showed that RNA can function as a catalysis. Slide5
Hairpin Ribozyme
Found
in plant viruses
Composed
of two helical domains containing a small and large internal loop.
These two domains must work together in order to form its active conformation. Slide6Slide7Slide8
Hammerhead Ribozyme
Name comes from shape of the secondary structure
“3d structure consists of three helices of variable length and a sequence of 11 unpaired nucleotides surrounding the cleavage site”.Slide9Slide10Slide11
RNase P
A
ribonuclease
that serves as a catalyst
Will often cleave RNA
Catalyzes the maturation of T-RNA Slide12Slide13Slide14
Future Applications
Specially
engineered ribozyme sheers could be used in
Antiviral medication
Production of plants with antivirus properties
Could potentially be used to correct genetic disordersSlide15
Multifactoral Catalysis of Ribozymes & Mechanism
S
N
2 reaction: nucleophilic
attack by the 2′-O on the 3′-P, with departure of 5′-O Slide16
Ribosome Function: Nucleolytic Cleavage
Transesterification
reactions
:Slide17
Ribozymes Function
Nucleotides in RNA can function to catalyze
reactions such as peptide bond formation
The
peptidyl
transferase center of cells (r
ibosome) is a
ribozymeSlide18
Ribozymes – pH & Metal Ions
RNA lacks nucleobase functional groups
pK
a
’s
– unlikely contributor to ribozyme chemistry. However…‘Metallozymes’
Strobel
and Cochrane 2007
Slide19
Acid-Base Catalysis of Nucleobases from RNA
A
(Adenine)38 functioning as an acid
(Guanine) 8 functioning as a base
Bevilacqua
, Biochemistry 2003Slide20
John K.
Frederiksen
a
,
1
and Joseph A.
Piccirilli 2009Slide21
Group I intron splicing promoted by two catalytic metal ions.
The
nucleophile
(U-1 O3′), scissile phosphate, leaving group (ΩG O3′), and labile bond
Catalytic metal ions
Strobel
and
Cochrane 2007
Slide22
Hairpin Ribozyme
Nucleobases
interact with
ribozyme
Transition state stabilized by
hydrogen bonds
Slide23
Hairpin Ribozyme
Transition state stabilized by H-bonds
Ruppert
et al, Science 2002Slide24
Kinetics of Ribozymes
Michaelis
-
Menton
Kinetics:
E + S ES E + P
Catalytic Efficiency: Ribozyme Km values comparable to Km values of protein enzymes Ribozymes KCAT are lower than those values observed for protein enzymes.
K
CAT
/K
M
= 10
3
-10
6
M
-1
.min
-1
10
11
times faster than
the un-catalyzed reaction