Intrasectoral Zoning of Proteins and Nucleotides in Simple Crystalline Hosts
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Intrasectoral Zoning of Proteins and Nucleotides in Simple Crystalline Hosts
Miki Kurimoto, Loyd D. Bastin, Daniel Fredrickson, Pamela N. Gustafson, Sei-Hum Jang, Werner Kaminsky, Scott Lovell, Christine A. Mitchell, Jean Chmielewski,1 Bart Kahr Department of Chemistry, Box 351700, University of Washington, Seattle, WA 98195-1700, U.S.A. 1 Department of Chemistry, 1393 Brown Laboratories, Purdue University, West Lafayette, IN 47907-1393, U.S.A. ABSTRACT Oriented gases of biopolymers in simple, single crystal hosts might be used to measure anisotropic molecular properties of analytes that could not otherwise be crystallized. Here we show two types of crystals as examples of the single crystal matrix isolation of biopolymers: green fluorescent protein in α-lactose monohydrate as a model system for studying the kinetic stabilization of biopharmaceuticals, and adenosine phosphates in potassium dihydrogen phosphate, a first step in the matrix isolation of oligonucleotides. In each case, the hosts undergo compositional zoning – both intersectoral and intrasectoral – during growth from solution. Intrasectoral zoning is evident by the selective luminescence of adjacent vicinal slopes of growth active hillocks. Nucleotides furthermore distinguish between symmetry related growth sectors enantioselectively.
INTRODUCTION Efforts to understand how and why large guest molecules enter simple carboxylic acid host crystals during growth from solution have taken on a new urgency in light of the explosive growth of matrix-assisted laser desorption ionization mass spectrometry (MALDI-MS), a revolutionary process in which biopolymer analytes precipitated with a crystalline acid matrix, are propelled into the gas phase upon irradiation [1]. The success of the MALDI experiment depends critically on the choice of host crystal but the characteristics of a good host such as 2,5dihydroxybenzoic acid, or of a poor one such as the constitutional isomer 3,5-dihydroxybenzoic acid, are still not understood. The inventors of MALDI-MS demonstrated that when large crystals of matrices were grown in the presence of proteins, cleaved, and irradiated on the fresh surface, the signal in the mass spectrum was no less intense than that from the irradiation of a polycrystalline precipitate, suggesting that the growth of genuine mixed crystals is necessary for the transfer of energy from the laser to matrix to analyte [2]. This question has been studied intensively since 1991, but a current review of the subject is equivocal: “Protein incorporation into the crystals of solid MALDI matrices is helpful, but not a prerequisite [3].” We demonstrated, however, that when single crystals of sinapic acid (3,5-dimethoxy-4hydroxy-trans-cinnamic acid) grown in the presence of myoglobin were analyzed by MALDIMS, the intensity of the protein signal in the mass spectrum was highly dependent upon the region of the crystal irradiated with the focused laser. This was a consequence of intersectoral zoning, the compositional partitioning of impurities in crystals between or
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