Garman lab: Interconversion of lysosomal enzyme specificities: Difference between revisions

The goal of this page is to provide three-dimensional and interactive figures to explore the structures determined for the 2010 paper "Interconversion of the Specificities of Human Lysosomal Enzymes Associated with Fabry and Schindler Diseases" by Ivan B. Tomasic, Matthew C. Metcalf, Abigail I. Guce, Nathaniel E. Clark and Scott C. Garman<ref name=primary>DOI: 10.1074/jbc.M110.118588</ref>. The starting point are the figures found in this paper. Biochemistry students at Westfield State University recreated these figures in jmol, and revised them after getting feedback from the authors. A special thank you goes to Susan Al Mahrwuth, Samuel J. Butler, Susy Civil, Westin G. Cohen, Allison F. DeVivo, Tyler S. Fassett, Courtney M. Fisher, Kimberly Garcia, Stephanie L. Hardy, Maureen W. Kamau, Sienna R. Kardel, Allyson L. Kress, Julia M. Lahaie, Stephen A. Malerba, Brittany E. Ricci, Kimberly Rosario, Yelena Vynar, and Deanna N. Womack for creating the initial figures and captions. If you are interested to learn how these figures were made, take a look at the discussion page (2nd tab above).

The research is about two related enzymes, GAL and NAGAL. They are found in the same location in the human body (in the lysosome, an acidic organelle responsible for breaking down molecules the cell no longer needs), their primary sequence is 50% identical, they catalyze the same reaction (hydrolysis of α-glycosidic bonds), and they have very similar active sites. However, they differ in substrate specificity (one cleaves the bond with galactose, and the other with N-acetyl glalactosamine). In terms of structure, the backbone conformation is quite similar (Fig. 1 panel B and C: compare <scene name='78/786673/Gal_overall/5'>GAL</scene> and <jmol>

<jmolLink>

  <script> model 2

  </script>

  <text>NAGAL</text>

</jmolLink>

</jmol>). The active sites differ in only two residues, and the substrates bind in almost the same way (Fig 1 panel B and C insets: compare the active site of <scene name='78/786673/Gal_active_site/5'>GAL</scene> and <jmol>

  <script> model 2

  </script>

  <text>NAGAL</text>

</jmolLink>

</jmol>).  

To test this, they made variants called GAL(SA) and NAGAL(EL), in which one active site has the amino acids of the other active site and vice versa (by swapping the two residues that are different). The data obtained by enzyme kinetics supported their hypothesis; the preference for galactose vs N-acetyl galactosamine is swapped (not shown here, but the data is in their paper<ref name=primary/>). Crystal structures (Fig. 2 panel A and B) show how GAL(SA) is able to bind to either <scene name='78/786673/Fig2a_galnac_complex/2'>N-acetyl galactosamine</scene> or <scene name='78/786673/Galsa_gal/21'>galactose</scene>. Comparing the structures of the NAGAL: N-acetyl galactosamine complex and the GAL(SA): N-acetyl galactosamine complex (Fig. 2 panel D) shows that <scene name='78/786673/Galsa_nagal/2'>they bind the ligand in a very similar manner</scene>.

<script>!exit; ppdiaCaptionCmd = "changeCaption('The enyme GAL (green) in complex with the sugar galactose (thick wireframe), with glycolysation shown as thin wireframe. View along the 2-fold axis of the dimer. ','white','black');"; javascript @ppdiaCaptionCmd; delay 1.2; model 1</script>

</jmol><jmol>

<script>!exit; ppdiaCaptionCmd = "changeCaption('The enyme NAGAL (blue) in complex with the sugar N-acetyl galactosamin (thick wireframe), with glycolysation shown as thin wireframe. View along the 2-fold axis of the dimer. ','white','black');";

   javascript @ppdiaCaptionCmd;delay 1.5; model 2</script>

</jmol>

<jmol>

<script>!exit; ppdiaCaptionCmd = "changeCaption('Superposition of GAL (green) and NAGAL (blue). View along the 2-fold axis of the dimer. ','white','black');"; javascript @ppdiaCaptionCmd; delay 1.5; model 0</script>

</jmol>

<jmol>

<script>animation mode loop; ppdiaCaptionCmd = "changeCaption('Superposition of GAL (green) and NAGAL (blue). View along the 2-fold axis of the dimer. ','white','black');"; javascript @ppdiaCaptionCmd; animation on</script>

<scene name='78/786673/Bonus/1'>α-GAL active site</scene>

<scene name='78/786673/Fig2a_galnac_complex/2'>Panel A</scene>: in complex with N-acetyl galactosamine

<scene name='78/786673/Galsa_gal/21'>Panel B</scene>: in complex with galactose

Superposition of structures: GAL(SA) bound to N-acetyl glucosamine, in yellow, and NAGAL bound to N-acetyl glucosamine, in dark blue. The sugar bound to GAL(SA) is shown in light brown and the one bound to NAGAL is shown in light blue.

<jmolButton>

<script>animation mode loop; animation on</script>

  <text>animate</text>

</jmolButton>