Modular proteins II

1
Modular proteins II

• Level 3 Molecular Evolution and Bioinformatics
• Jim Provan

Patthy Sections 8.1.3 8.2
2
Intron phase
3
Intron phase and evolution of collagenases
4
Exon shuffling by intronic recombination

• Middle repetitive sequences flanking an exon may
  facilitate looping out or insertion of modules
  by intronic recombination
• Best example of contraction and expansion of a
  multidomain protein found in apolipoprotein(a)
• Number of tandem kringle domains ranges from 12
  to 51 copies
• In one variant, 24 of the 37 kringle domains have
  identical nucleotide sequences, suggesting very
  recent duplication
• Isoforms containing different numbers of kringle
  domains do not follow simple Mendelian patterns
  of inheritance offspring often have
  apolipoprotein(a) isoforms that differ from those
  of parents
• Such proteins retain interdomain introns these
  are responsible for high levels of gene structure
  plasticity

5
Factors favouring intronic recombination

• Only a tiny portion of spliceosomal introns is
  essential for splicing
• 5 end, 3 end and branch site
• Separated by very long sequences that are
  tolerant to insertions and deletions
• Illustrated by comparison of urokinase genes
• Genome organisation of murine, human, porcine and
  chicken genes is identical in terms of location
  and phase class of introns
• Chicken urokinase introns show hardly any
  sequence similarity with corresponding mammalian
  introns, except near splicing junctions and
  branch sites
• Great difference in size of orthologous introns
• Intron A is 1489 bp in chicken
• Only 306 bp in humans

6
Factors favouring intronic recombination

• Advantages of spliceosomal introns for exon
  shuffling (large size, presence of middle
  repetitive sequences, tolerance to structural
  changes) holds primarily for vertebrate genomes
• Fungi and plants have fewer and shorter introns
• Genes of best studied invertebrate genomes (C.
  elegans, D. melanogaster) also have shorter
  introns
• Relatively compact genome may be characteristic
  of ancestral metazoa
• Alternatively, selection may have led to a
  secondary increase in genome compactness in these
  lineages
• Since plant spliceosomal introns are shorter than
  those of vertebrates, they are less suitable for
  intronic recombination
• Splicing of chimeric introns, an inevitable
  consequence of intronic recombination, is
  impaired in yeast and plants

7
Acceptance of mutants created by intronic
recombination

• Several levels of selection determine whether
  intronic recombination mutant will be fixed or
  rejected
• Chimeric intron must be spliced correctly,
  otherwise translation will probably run into a
  stop codon in the mRNA/intron region and form a
  truncated protein
• Two non-orthologous introns must be in the same
  phase class
• Must split the reading frame in the same phase
• Downstream exon must be translated in its
  original phase to prevent frameshift mutations
• Symmetrical exons
• New protein must be able to adopt a stable
  conformation
• Selective advantage of having a new functional
  domain
• Impact of exon insertion may initially be
  mitigated by alternate splicing

8
The intron-phase compatibilty rule
9
The symmetrical exon rule

• Insertion, deletion and duplication of a module
  by intronic recombination can satisfy the phase
  compatibility requirement only if the two introns
  flanking the module are of the same phase
  (symmetrical modules)
• Only symmetrical module groups are 0-0, 1-1 and
  2-2
• Can only be inserted into the compatible intron
  i.e. 1-1 modules can only be inserted into phase
  1 introns
• If the structure of a gene of a modular protein
  conforms to these rules, it suggests that the
  protein has evolved through exon shuffling

10
Intron insertion and removal
11
Evolution of mobile modules

• Conversion of a domain to a module
• Protein domain may be converted to a protomodule
  if introns of identical phase are inserted at its
  boundaries
• Tandem duplication may lead to homopolymerisation
• New mobile module may be excised and reinserted
  at a new location
• There is a large variety of class 1-1 modules
  known, but relatively few class 2-2 or class 0-0
  modules
• May be due to initial predominance of class 1-1
  modules by chance

12
Conversion of a domain to a mobile module
13
Example of the modularisation process

• The Kunitz type proteinase inhibitor is a single
  module protein
• In bovine pancreatic trypsin inhibitor gene,
  phase 1 intons are found at both boundaries of
  the single inhibitor domain (protomodule stage)
• Lipoprotein-associated coagulation inhibitor
  consists of three tandem copies of this module
  (tandem duplication stage) each module is
  encoded by a distinct class 1-1 exon
• Kunitz-type inhibitor modules have been inseted
  into the genes of other proteins (shuffling
  stage)
• Amyloid precursos
• Collagens

14
Evolution of exon shuffling

• Obvious examples of proteins assembled by exon
  shuffling are restricted to animals
• Not surprising, considering that the evolution of
  introns and modules is a relatively late
  development
• Recent evolution of spliceosomal pre-mRNA
• Large-scale genome projects on model organisms
  provides information on modular evolution
• Many examples in metazoa presence of
  vertebrate modules in invertebrates suggests
  mechanism predates split
• No evidence in yeast
• Only one possible example in Arabidopsis
  (receptor protein kinase with two EGF-like
  domains)

15
Evolutionary significance of exon shuffling

• Number of proteins constructed from modules
  underlines value of exon shuffling
• Several unique features made this mechanism
  important
• Large collection of binding specificities can
  coexist in a single protein e.g. plasma
  proteinases
• Acquisition of a new domain can bring about a
  sudden change in specificity
• Good example is gelatinase
• Insertion of a gelatin-binding FN2 module into an
  ancestral metalloproteinase of the collagen
  family
• Could be correlated with metazoan big bang

16
Modular assembly by exonic recombination

• Exon shuffling by intronic recombination may not
  be the only way to exchange domains between
  genes
• Modular protein of the bacteria
  Peptostreptococcus magnus is the product of a
  recent intergenic recombination of two different
  types of streptococcal surface proteins
• Transfer of one part of a prokaryotic gene to
  another without the aid of introns
• Multidomain bacterial proteins of the PEP sugar
  transferase system
• Three functional domains separated by unusual
  flexible linker regions
• Linkers responsible for frequent rearrangements