Conformation of Highly-Charged Gas-Phase Lysozyme Revealed by Energetic Surface Imprinting

We present new results from an energetic surface imprinting method which allows us to outline the general conformation of protein ions in vacuo. Both disulfide-bond-intact and disulfide-bond-reduced gasphase lysozyme ions were produced by electrospray ionization and were accelerated and impacted onto graphite surfaces. The resulting surface defects, each created by a single incident ion, were imaged with scanning force microscopy. Disulfide-intact lysozyme ions created compact, slightly elliptical hillocks on the surfaces, whereas disulfide-reduced lysozyme produced more oblong, elongated hillocks. By employing a thermal model describing the response of graphite to energy deposited by an elongated incident energetic projectile, we calculated from the hillock sizes for disulfide-reduced lysozyme (Q = 14+) an overall length of 32.1 +/- 1.6 nm. This value is close to the length we observe for apomyoglobin (Q = 14+), 35.5 +/- 2.4 nm, although apomyoglobin and lysozyme possess significantly different numbers of amino acid residues. Based on these results, we hypothesize that aspects of a protein's native secondary structure are preserved in the gas phase, even if the tertiary structure might be non-native. We have unfolded disulfide-intact lysozyme computationally and find a qualitatively good agreement with the experimentally obtained length of disulfide-intact (Q = 9+) lysozyme.