Difference between revisions of "Protein domains/Overview"
 
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Protein tags and modifiers are short peptide sequences cloned in frame with protein coding sequences that change the protein's behavior.
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Protein domains encode portions of proteins and can be assembled together to form translational units, a genetic part spanning from translational initiation (the RBS) to translational termination (the stop codon).
  
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Protein tags and modifiers might
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[[Image:ProteinDomains.png|center]]
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*change the protein's location (localization signals)
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*enable it to be readily purified (affinity tags)
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*alter the protein's degradation rate (degradation tags)
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*target the protein for cleavage (cleavage sites)
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Protein tags may include a start codon or a stop codon, but not both, since they are intended to be assembled in frame with protein coding sequences.
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There are several different types of protein domains.
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#'''Head Domain''': The Head domain consists of the start codon followed immediately by zero or more triplets specifiying an N-terminal tag, such as a protein export tag or lipoprotein binding tag.  Head domains should begin with an <code>ATG</code> start codon and include codons 2 and 3 of the protein at a minimum.  Examples of head domains include
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#*<code>ATG</code> start codon
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#*<code>ATG</code> start codon and codons 2-3
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#*<code>ATG</code> start codon and signal sequence
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#*<code>ATG</code> start codon and affinity tag
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#'''Internal Domains''': Protein domains consist of a series of codon triplets coding for an amino acid sequence without a start codon or stop codon.  Multiple Internal Domains can be fused.  Examples of internal domains include
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#*DNA binding domains
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#*Dimerization domains
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#*Kinase domains
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#'''Special Internal Domains''': Short Domains with specific function may be separately categorized, but obey the same composition rules as normal internal domains.  Examples of special internal domains include
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#*Linkers
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#*Cleavage sites
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#*Inteins
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#'''Tail Domain''': The C-terminus of a coding region consists of zero or more triplet codons, followed by a pair of TAA stop codons.  In the simplest case, the stop codons terminate the protein with an Stop.  More complex Tails may include degradation tags appropriate to the organism (i.e., with different degradation rates). Examples of Tail domain include
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#*<code>TAATAA</code> stop codons
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#*A degradation tag followed by <code>TAATAA</code> stop codons
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#*An affinity tag followed by <code>TAATAA</code> stop codon
  
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'''You may also be interested in'''
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Unfortunately, the original BioBrick assembly standard, Assembly standard 10, does not support in-frame assembly of protein domains.  (Assembly standard 10 creates an 8 bp scar between adjacent parts.)  Therefore, it is recommended that you use an alternate approach to assemble protein domains together to make a translational unit.  There are several possible approaches to [[Help:Protein domains/Assembly|assembling protein domains]] including direct synthesis (preferred because it creates no scars) as well as various assembly standards.  Regardless of which standard you choose, we suggest that the resulting protein coding sequence or translational unit comply with the [[Help:Assembly standard 10|original BioBrick assembly standard]] so that your parts can be assembled with most of the parts in the Registry.
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{|
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|valign='top' align='center' width=50px | {{Click || image=Part icon cds.png | link=Protein coding sequences |width=30px | height=30px}}
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|valign='top' |'''[[Protein coding sequences]] [[Help:Protein coding sequences|(?)]]''': Protein coding sequences encode the amino acid sequence of a particular protein.  Note that some protein coding sequences only encode a protein domain or half a protein. Others encode a full-length protein from start codon to stop codon.
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''Protein coding sequences should be as follows''
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|}
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<font face="courier" size="3">'''GAATTC GCGGCCGC T TCTAG <font color="red">[ATG ... TAA TAA]</font> T ACTAGT A GCGGCCG CTGCAG'''</font>
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Note: Although most RBSs are currently specified as separate parts in the Registry, we are now moving to a new design in which the RBS and Head domain are combined into a single part termed a '''Translational start'''.  The new design has the advantage of encapsulating both ribosome binding and translational initiation within a single part.  Our working hypothesis is that the new design will reduce the likelihood of unexpected functional composition problems between the RBS and coding sequence.

Latest revision as of 22:57, 8 December 2009

Protein domains encode portions of proteins and can be assembled together to form translational units, a genetic part spanning from translational initiation (the RBS) to translational termination (the stop codon).

ProteinDomains.png

There are several different types of protein domains.

  1. Head Domain: The Head domain consists of the start codon followed immediately by zero or more triplets specifiying an N-terminal tag, such as a protein export tag or lipoprotein binding tag. Head domains should begin with an ATG start codon and include codons 2 and 3 of the protein at a minimum. Examples of head domains include
    • ATG start codon
    • ATG start codon and codons 2-3
    • ATG start codon and signal sequence
    • ATG start codon and affinity tag
  2. Internal Domains: Protein domains consist of a series of codon triplets coding for an amino acid sequence without a start codon or stop codon. Multiple Internal Domains can be fused. Examples of internal domains include
    • DNA binding domains
    • Dimerization domains
    • Kinase domains
  3. Special Internal Domains: Short Domains with specific function may be separately categorized, but obey the same composition rules as normal internal domains. Examples of special internal domains include
    • Linkers
    • Cleavage sites
    • Inteins
  4. Tail Domain: The C-terminus of a coding region consists of zero or more triplet codons, followed by a pair of TAA stop codons. In the simplest case, the stop codons terminate the protein with an Stop. More complex Tails may include degradation tags appropriate to the organism (i.e., with different degradation rates). Examples of Tail domain include
    • TAATAA stop codons
    • A degradation tag followed by TAATAA stop codons
    • An affinity tag followed by TAATAA stop codon

Unfortunately, the original BioBrick assembly standard, Assembly standard 10, does not support in-frame assembly of protein domains. (Assembly standard 10 creates an 8 bp scar between adjacent parts.) Therefore, it is recommended that you use an alternate approach to assemble protein domains together to make a translational unit. There are several possible approaches to assembling protein domains including direct synthesis (preferred because it creates no scars) as well as various assembly standards. Regardless of which standard you choose, we suggest that the resulting protein coding sequence or translational unit comply with the original BioBrick assembly standard so that your parts can be assembled with most of the parts in the Registry.


Protein coding sequences should be as follows

GAATTC GCGGCCGC T TCTAG [ATG ... TAA TAA] T ACTAGT A GCGGCCG CTGCAG


Note: Although most RBSs are currently specified as separate parts in the Registry, we are now moving to a new design in which the RBS and Head domain are combined into a single part termed a Translational start. The new design has the advantage of encapsulating both ribosome binding and translational initiation within a single part. Our working hypothesis is that the new design will reduce the likelihood of unexpected functional composition problems between the RBS and coding sequence.