Difference between revisions of "ABC-INT-Homology modelling"

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This is a "milestone unit". Its purpose is merely to collect a number of preparatory units into a single, common prerequisite. It has no contents of its own; you are expected to be familiar and competent with all preparatory material at this point.
 
This is a "milestone unit". Its purpose is merely to collect a number of preparatory units into a single, common prerequisite. It has no contents of its own; you are expected to be familiar and competent with all preparatory material at this point.
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=== The DNA binding site ===
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Now, that you know how YFO Mbp1 aligns with yeast Mbp1, you can evaluate functional conservation in these homologous proteins. You probably already downloaded the two Biochemistry papers by Taylor et al. (2000) and by Deleeuw et al. (2008) that we encountered in Assignment 2. These discuss the residues involved in DNA binding<ref>([http://www.ncbi.nlm.nih.gov/pubmed/10747782 Taylor ''et al.'' (2000) ''Biochemistry'' '''39''': 3943-3954] and [http://www.ncbi.nlm.nih.gov/pubmed/18491920 Deleeuw ''et al.'' (2008) Biochemistry. '''47''':6378-6385])</ref>. In particular the residues between 50-74 have been proposed to comprise the DNA recognition domain.
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{{task|
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# Using the APSES domain alignment you have just constructed, find the YFO Mbp1 residues that correspond to the range 50-74 in yeast.
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# Note whether the sequences are especially highly conserved in this region.
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# Using Chimera, look at the region. Use the sequence window '''to make sure''' that the sequence numbering between the paper and the PDB file are the same (they are often not identical!). Then select the residues - the proposed recognition domain -  and color them differently for emphasis. Study this in stereo to get a sense of the spatial relationships. Check where the conserved residues are.
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# A good representation is '''stick''' - but other representations that include sidechains will also serve well.
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# Calculate a solvent accessible surface of the protein in a separate representation and make it transparent.
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# You could  combine three representations: (1) the backbone (in '''ribbon view'''), (2) the sidechains of residues that presumably contact DNA, distinctly colored, and (3) a transparent surface of the entire protein. This image should show whether residues annotated as DNA binding form a contiguous binding interface.
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DNA binding interfaces are expected to comprise a number of positively charged amino acids, that might form salt-bridges with the phosphate backbone.
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{{task|
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*Study and consider whether this is the case here and which residues might be included.
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Revision as of 04:26, 31 August 2017

Integration Unit: Homology Modelling


 

Keywords:  Integration unit: create a homology model and assess the role of sequence conservation


 



 


Sorry!

This page is only a stub; it is here as a placeholder to establish the logical framework of the site but there is no significant content as yet. Do not work with this material until it is updated to "live" status.


 


Abstract

This page assesses the learning units for working with multiple sequence alignments and structure data.

  • Use an Mbp1 orthologue (RBM)
  • Find all orthologues in YFO and reference species
  • Homology model
  • Calculate conservation scores
  • Map, visualize
  • Interpret


 


This unit ...

Prerequisites

You need to complete the following units before beginning this one:


 


Objectives

...


 


Outcomes

...


 


Deliverables

  • No separate deliverables: This unit collects other units and has no deliverables on its own.


 


Evaluation

Evaluation: NA

This unit is not evaluated for course marks.


 


Contents

This is a "milestone unit". Its purpose is merely to collect a number of preparatory units into a single, common prerequisite. It has no contents of its own; you are expected to be familiar and competent with all preparatory material at this point.


The DNA binding site

Now, that you know how YFO Mbp1 aligns with yeast Mbp1, you can evaluate functional conservation in these homologous proteins. You probably already downloaded the two Biochemistry papers by Taylor et al. (2000) and by Deleeuw et al. (2008) that we encountered in Assignment 2. These discuss the residues involved in DNA binding[1]. In particular the residues between 50-74 have been proposed to comprise the DNA recognition domain.

Task:

  1. Using the APSES domain alignment you have just constructed, find the YFO Mbp1 residues that correspond to the range 50-74 in yeast.
  2. Note whether the sequences are especially highly conserved in this region.
  3. Using Chimera, look at the region. Use the sequence window to make sure that the sequence numbering between the paper and the PDB file are the same (they are often not identical!). Then select the residues - the proposed recognition domain - and color them differently for emphasis. Study this in stereo to get a sense of the spatial relationships. Check where the conserved residues are.
  4. A good representation is stick - but other representations that include sidechains will also serve well.
  5. Calculate a solvent accessible surface of the protein in a separate representation and make it transparent.
  6. You could combine three representations: (1) the backbone (in ribbon view), (2) the sidechains of residues that presumably contact DNA, distinctly colored, and (3) a transparent surface of the entire protein. This image should show whether residues annotated as DNA binding form a contiguous binding interface.


DNA binding interfaces are expected to comprise a number of positively charged amino acids, that might form salt-bridges with the phosphate backbone.


Task:

  • Study and consider whether this is the case here and which residues might be included.




 


Further reading, links and resources

 


Notes

  1. ↑ (Taylor et al. (2000) Biochemistry 39: 3943-3954 and Deleeuw et al. (2008) Biochemistry. 47:6378-6385)


 


Self-evaluation

 



 



 

If in doubt, ask! If anything about this learning unit is not clear to you, do not proceed blindly but ask for clarification. Post your question on the course mailing list: others are likely to have similar problems. Or send an email to your instructor.


 

About ...
 
Author:

Boris Steipe <boris.steipe@utoronto.ca>

Created:

2017-08-05

Modified:

2017-08-09

Version:

0.1

Version history:

  • 0.1 First stub

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