1ENH and 1ZTR

1ENH: Native Drosophila melanogaster engrailed homeodomain
1ZTR: Denatured Drosophila melanogaster engrailed homeodomain mutant L16A

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I. Introduction

1ENH, from the fruit fly Drosophila melanogaster, is a 61-residue, 3-alpha helix bundle protein (H1, H2, H3).

A novel mutant of 1ENH, given the code 1ZTR, directly solves the solution structure of a true denatured state under conditions that would typically favor folding. 

Denaturation had been encouraged by a L16A mutation in 1ENH.  L16A is a fully buried residue located in the middle of H1.  The mutation from leucine to alanine changed the side chain from a larger hydrophobe to a smaller one, which decreased van der Waals interactions and decreased protein stability.

Changes in ionic strength cause 1ZTR to fold and unfold reversibly.  At high ionic strengths, the mutant protein exists in its folded, native state, but at physiological ionic strength       (~ 150 mM) the protein denatures. 

The solution structure of 1ZTR was obtained by 1HNMR.  The above structure of 1ZTR shows a superimposition of the 25 lowest-energy conformer structures.  Only one of these conformer structures was selected for display in this Jmol.

For more information see Biochemistry Online: Chapter 2D - PROTEIN FOLDING/STABILITY IN VIVO AND IN VITRO

II. General Structure

Native State (1ENH)

Spacefill
Backbone

The native state of En-HD is compact in shape

Rocket
H Bonds
Hydrophobes

Hydrophobic residues are mostly in the interior of the protein

Denatured State (1ZTR)

Spacefill

Because this structure was determined by 1HNMR, hydrogens show up (as white)

Backbone

The denatured state is larger, more elongated, and less compact than the native state

Rocket

H1 is longer by at least one turn compared to the native state

H Bonds

While the core is disrupted in the denatured state, the helical structure and the turn connecting H2 and H3 remains intact

Hydrophobes

Numerous hydrophobic residues are exposed at the surface

Superimposition of 1ENH and 1ZTR

Native state shown in light blue, denatured state shown in purple

Spacefill
Backbone
Rocket
H Bonds

Religa, T. L., Markson, J. S., Mayor, U., Freund, S. M. V., Fersht, A. R. Solution structure of a protein denatured state and folding intermediate. Nature 437, 1053-1056 (13 October 2005).