Modular proteins I

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Modular proteins I

• Level 3 Molecular Evolution and Bioinformatics
• Jim Provan

Patthy Sections 8.1.1 8.1.3
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Protein domains

• Folded structures of proteins that are larger
  than ?200-300 residues generally consist of
  multiple structural domains
• Compact, stable units with a unique
  three-dimensional structure
• Interactions within a domain are more significant
  than those between domains
• Fold independently i.e. structural domains are
  also folding domains
• If domain performs distinct function which
  remains intact in the isolated domain, then it is
  also a functional domain
• Many multidomain proteins are homomultimeric i.e.
  contain multiple copies of a single type of
  structural domain
• Arisen through internal duplication of complete
  domains
• Fate of domains determined by similar rules to
  paralogous genes

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Protein domains

• Many multidomain proteins are heteromeric
• Example is plasminogen activator where a
  trypsin-like serine protease is joined to
  kringle, finger and EGF domains
• May occur by fusion of two or more genes
  (chimeric proteins)
• Also known as modular proteins, with domains
  known as modules
• Certain modules occur in a wide variety of
  hetero- and homomultimeric proteins
• Suggests mechanisms to facilitate duplication and
  dispersal
• Building blocks of different types of
  multidomain proteins are known as mobile protein
  modules
• Frequency of transfer and incorporation into new
  protein reflects fixation probability

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Modular assembly by intronic recombination

• Discovery of introns provided potential new
  mechanisms for protein evolution
• Gilbert suggested that recombination within
  introns could assort exons independently
• Idea of rapid construction of novel genes from
  parts of old ones led to the formulation of the
  exon-shuffling hypothesis
• According to introns early theories, all extant
  genes were constructed from a limited number of
  exon types
• Under the introns late theory, intronic
  recombination and exon shuffling could not have
  played a major role in the assembly of the
  earliest genes
• Original theory was that exons corresponded
  directly to modules and/or structural motifs

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Problems with the introns early hypothesis

• In the case of many genes, no obvious
  correspondence was observed between protein
  structure and intron location
• It is now known that introns can also be inserted
  into genes i.e. structure of a gene may not be
  its original structure
• Introns suitable for exon shuffling did not
  originate until a relatively late stage of
  eukaryotic evolution
• Exon shuffling has only been conclusively
  demonstrated in young proteins unique to higher
  eukaryotes
• Only a special group of exons, the symmetrical
  modules, are really valuable for exon shuffling.
  Intron phase distribution is also a crucial
  factor.

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Self-splicing introns

• Group I introns
• Reaction requires only a guanine nucleotide
  cofactor
• Provides a free 3-OH group that attacks the 5
  splice site
• 3-OH generated at the end of the upstream exon
• Second transesterification joins the two exons
• Crucially depends on folded structure of the
  intron itself
• Group II introns
• Does not require an external cofactor 2-OH of
  an adenine within the intron cuts the 5 splice
  site
• 25 phosphodiester bond (branch site) forms the
  lariat structure
• Although folding is still crucial, chemistry,
  sequence of events and lariat formation are
  similar to nuclear spliceosomal introns

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Spliceosomal intron splicing mechanism
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Spliceosomal introns

• Spliceosomal introns are only spliced in the
  presence of a complex of specific proteins and
  RNA known as a spliceosome
• Majority of intron is unimportant as long as the
  5 and 3 splice sites and the branch site are
  conserved, splicing can take place
• Large insertions into spliceosomal introns, or
  deletions do not affect splicing efficiency
• Chimeric introns, containing the 5 end of one
  intron and the 3 end of another, are also
  properly spliced
• Mutations (directed or otherwise) in these
  regions lead to aberrant splicing
• Spliceosomal intron plays a minor role in its own
  splicing the actual spliceosome complex is more
  important

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Evolution of spliceosomal introns

• Both group I and group II self-splicing
  mechanisms resemble spliceosome catalysed
  splicing
• Initial step is attack by ribose hydroxyl group
  on 5 splice site
• In each case, reactions are transesterifications
  where phosphate moieties are retained in products
• In group II and spliceosomal introns, intron is
  released as a lariat
• Accepted that spliceosomal-catalysed splicing
  evolved from group II self-splicing introns
• Key step was transfer of catalytic role from
  intron to other molecules
• Formation of spliceosome gave spleceosomal
  introns structural freedom as they no longer had
  to fulfil the catalytic function
• Generally found only in nuclear genomes of higher
  eukaryotes (plants, animals and fungi)

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Insertion and spread of spliceosomal introns
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Intron loss

• Plays a significant role in changing exon-intron
  structure of genes
• Introns may be eliminated through mechanism that
  gives rise to processed genes (retroposition)
• Reverse transcription can also lead to loss of
  only some introns
• Reverse transcription of perfectly spliced mRNA
  and recombination with the functional gene
  mutates original gene
• Partially processed pre-mRNA could give rise to a
  semi-processed gene generates a new paralogue

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Gene duplication / deletion due to intronic
recombination
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Exon shuffling via recombination in introns

• Believed that insertion of exons may occur by the
  same mechanism as insertion of introns
• Exon shuffling may be a consequence of the
  occasional inclusion of exon sequences in the
  insertion cycle of introns
• Alternative splicing (exon skipping during
  splicing) may yield exons with flanking introns
• If such a composite is inserted into the genome
  by the same mechanism that inserts single introns
  (reverse splicing) we have exon shuffling
• Key difference between intronic recombination
  model and retrotransposition model
• In first case, insertion occurs into a
  pre-existing intron of same phase as introns
  flanking exon
• Retrotransposition model does not have this
  requirement

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Evolution of urokinase
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Evolution of tissue plasminogen activator
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Evolution of Factor XII