joel 2:28, 29

©2000 Timothy G. Standish

Joel 2:28, 29 28 And it shall come to pass afterward,

that I will pour out my spirit upon all flesh; and your sons and your daughters shall prophesy, your old men shall dream dreams, your young men shall see visions:

29 And also upon the servants and upon the handmaids in those days will I pour out my spirit


RibozymesRibozymesTimothy G. Standish, Ph. D.


HistoryHistory At one time, in the not-too-distant past, only

proteins were thought to have catalytic properties Thomas Cech, working with Tetrahymena, wanted

to get a specific full-length mRNA, which consistently appeared on gels as multiple fragments

After working for over a year to get rid of the protein that “must” catalyze intron excision, Cech finally came to the conclusion that it was not contaminating protein, but the RNA itself that was doing the catalysis

For their discovery of catalytic RNA, Thomas Cech and Sydney Altman received the 1989 Nobel Prize in chemistry


Why So Excited?Why So Excited? Excitement about catalytic RNA revolved to a large

degree around the perception that somehow this eliminates the quandary of both proteins and nucleic acids being required for life

Perhaps a sort of proto-life once existed composed only of self-replicating RNAs which reached a certain level of sophistication and developed the ability to code for and produce proteins

DNA presumably entered into the picture somewhere along the way

In this model, smaller jumps are needed to get from non-living to living systems and catalytic RNAs may represent ancient remnants of life’s evolutionary history


A Disturbing RiddleA Disturbing Riddle“What makes the origin of life and of the genetic code a

disturbing riddle is this: the genetic code is without any biological function unless it is translated; that is, unless it leads to the synthesis of the proteins whose structure is laid down by the code. But, as Monod points out the machinery by which the cell (at least the nonprimitive cell which is the only one we know) translates the code ‘consists of a least fifty macromolecular components *which are themselves coded in DNA*’ (Monod, 1970; 1971, 143). Thus the code cannot be translated except by using certain products of its translation. This constitutes a really baffling circle: a vicious circle, it seems for any attempt to form a model, or a theory, of the genesis of the genetic code.”

Popper K., "Scientific Reduction and the Essential Incompleteness of All Science," in "Studies in the Philosophy of Biology," Vol. 259, 1974, p 259-284, p 270)


The RNA WorldThe RNA World“Once RNA is synthesized, it can make new copies of

itself only with a great deal of help from the scientist,” says Joyce of the Scripps Clinic, an RNA specialist. “It is an inept molecule,” he explains, “especially when compared with proteins.” Leslie E. Orgel of the Salk Institute for Biological Studies, who has probably done more research exploring the RNA-world scenario than any other scientist, concurs with Joyce. Experiments simulating the early stages of the RNA world are too complicated to represent plausible scenarios for the origin of life, Orgel says. “You have to get an awful lot of things right and nothing wrong,” he adds.

Horgan J. 1991. "In The Beginning...," Scientific American, February, p 103. Ellipses in original


Several Classes Of RNA Several Classes Of RNA Show Catalytic ActivityShow Catalytic Activity

1. Type I introns

2. Type II introns

3. E. coli RNAase P

4. Viroid and virusoid hammerhead ribozymes

5. Possibly some parts of ribozymes Most, if not all ribozymes are associated with proteins

in vivo and these may speed up and play other roles in the reaction

By themselves ribozymes are typically much slower than protein enzymes

Most require divalent cat ions to function


The Hammerhead RibozymeThe Hammerhead Ribozyme

AA

G

N’ N’ G

N CG

A

NN’ N’

N N N N

N’N’

GU

A

C

N

HUG

ACN

C

G

N

CleavagesiteCleavagesite

Stem IIStem II

Stem IIIStem III

Stem IStem I

Nucleotides shown in yellow are essential for efficient cleavage.

H = A, C or T




Stem IIStem II

Stem IIIStem III

Stem IStem I

Nucleotides shown in yellow and orange are essential for efficient cleavage.H = A, C or T

AA

N CG

NN N N N

HUG

ACN

G

N

G

N’ N’ GA

N’ N’ N’N’

GU

A

C

N

C



AA

G

N’ N’ G

N CG

A

NN’ N’

N N N N

N’N’

GU

A

C

N

HUG

ACN

C

G

N

Stem IIStem II

Stem IIIStem III

Stem IStem I


UC


AAAA

G

N’ N’ G

N CG

A

N

NN NN

N’ N’ N’ N’

NNN

GAN

HHC

G

UUGG

AACCNN NN Stem IIIStem IIIStem IIIStem III Stem IStem I


GUCC

N

UG

ACN N

N N

N’N’ N’N’ N’ N’

NN

A

A H

A

G

GA

N’

N’G

NC

NCG

Stem IIStem II

Stem IIIStem III Stem IStem I

Mg++


C

C


GUCC

N

UG

ACN N

N N

N’N’ N’N’ N’ N’

NN

A

A H

A

G

GA

N’

N’G

NC

NCG

Stem IIStem II

Stem IIIStem III Stem IStem I

Mg++



GUC

NA

GAG

NNNNN’N’

N’N’N’N’

N’N’

NN

NNHH

UG

ACN N

AAG

N’

N’NC

NCG

Stem IIStem II

Stem IIIStem III

Stem IStem IMg++




3D Structure3D Structure


The RNA Editing:The RNA Editing:A Heresy of Molecular Biology?A Heresy of Molecular Biology?

The central dogma of molecular genetics says that DNA is transcribed to make RNA which is translated to make proteins

Implicit in this dogma is the assumption that information flows in one direction, from DNA to RNA and then proteins, thus all information must originate directly in the sequence of bases in DNA

RNA editing provides a startling exception to this system

RNA editing involves modification, addition or deletion of bases resulting in changes in codon meaning


5’__ACG_GG___A___GA_____G_G____A__ACG__G_A_CCAG_A__G______

A_GG_____A_G_AGTTGAG___G____A___A_GGCG______GTTTTG_A__A__

_GG___A_G___A_____G_G__G_GAG___GCTTTCG______G___ACC__A_A_

G____G__G___A__A_G_GA__A_GG____G______A__GGTTA______AGA___

A___AA___G__GA_AAA_ACA____ATTTGT__G__AG_GG___A___G__AA___

____G____G_GTTTTTGG___AGG_______G__GTTTTG__G____G_A__A_GA

__GAG___G__G___GTTTTG______G_____G_GAAACCAG__ATGAGAGTTT

TTGCA__G__A___A__ACA__AAG__G_GGTGTTTT3’

5’ACGGGAGAGGAACGGACCAGAGAGGAGAGTTGAGGAAGGCGGTTTT

GAAGGAGAGGGGAGGCTTTCGGACCAAGGGAAGGAAGGGAGGTTAAG

AAAAGGAAAAACAATTTGTGAGGGAGAAGGGTTTTTGGAGGGGTTTTGG

GAAGAGAGGGGTTTTGGGGAAACCAGATGAGAGTTTTTGCAGAAACAA

AGGGGTGTTTT3’

5’UUACGUGGUUUAUUUGAUUUUUGUGUUUUAUUACGUUGUAUCCAGUAU

UGUUUUUUAUGGUUUUUAUGUAGU=GAGUUUGUUUUAUUUAUGGCGUU

UUUUGUU==GUAUUAUUUGGUUUAUGUUUAUUUUUGUGUUGUGAGUUU

GCUUUCGUUUUUUGUUUACCUUAUAUGUUUUGUUGUUUAUUAUGUGAUU

AUGGUUUUGUUUUUUAUUGGU=AUUUUUUAGAUUUAUUUAAUUUGUUGA

UAAAUACAUUUUAUUUGUUUGUUAGUGGUUUAUUUGUUAAUUUUUUUGU

UUUGUGUUUUUGGUUUAGGUUUUUUUGUUGUU==GUUGUUUUGUAUUA

UGAUUGAGUUUGUUGUUUG====GUUUUUUGUUUUUGUGAAACCAGUU

AUGAGAGUUU==GCAUUGUUAUUUAUUACAUUAAGUUGUGGUGUU==3’

Trypanosoma bruciTrypanosoma bruci Cytochrome C Oxidase IIICytochrome C Oxidase III

ACGT = DNA sequence ACGU = mRNA coded in DNA U = Bases inserted into mRNA T = Bases removed from mRNA

joel 2:28, 29

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