Ph.D. 1990, Weizmann Institute of Science; Lecturer, 1996

E-mail: irinak@macbeth.ls.huji.ac.il

Tel: 972-2-658-5575/62; Fax: 972-2-658-5573

Research Interests:

Structure determination of biological macromolecules and correlating their three-dimensional structure to their biological activity using multidimensional heteronuclear nuclear magnetic resonance spectroscopy (NMR) in solution.

Studying the dynamics of macromolecules in solution.

Developing new methods for structure determination and for studying dynamics.

Background on Structural NMR:

Over the past decade high-resolution multidimensional NMR spectroscopy has become a powerful method for determining the three dimensional structures of proteins and nucleic acids in solution. Using NMR spectroscopy one can investigate proteins in their "native state", at physiological conditions. Nuclear magnetic resonance is a spectroscopic method that uses the frequency of the nuclear spin to glean information about the chemical environment of the nucleus. Structural characteristics are directly obtained from two basic magnetic interactions: Homo- and heteronuclear J couplings and nuclear Overhauser effect (NOE). J splitting originates from scalar through-bond electron mediated spin-spin interactions between neighboring nuclei, providing information about torsion angles between the adjacent residues along the polypeptide backbone. The most important source of structural information stems from the fact that protons continuously exchange magnetizations with one another via through-space dipole-dipole interactions, the nuclear Overhauser effect. Measuring the intensity of the cross peaks in the NOESY spectrum yield semiquantitative values for the interproton distances in the macromolecule. These, together with the angular constraints based on coupling constants, are then used as restraints in distance geometry and restrained molecular dynamics calculations, to generate NMR-derived three dimensional structures.

Larger proteins of up to a molecular weight of approximately 50KD demand more complicated NMR methods since the signals overlap and little information can be directly determined from the aforementioned methods. In the case of larger bio-molecules, 15N and 13C enrichment is used where the 15N and/or 13C labels significantly increase the ability to separate the overlapped signals in the multidimensional spectra.

Once the NMR-derived structures are determined, structural analysis in terms of hydrophobicity, electrostatics and additional structural elements can begin. These are correlated to the biological activities of the molecules and used to elucidate mechanisms and enhance various characteristics by designed synthesis.

The NMR Facility:

My lab, the NMR laboratory, is at the Institute of Life Sciences at the Hebrew University Givat Ram campus. The laboratory hosts a state-of-the-art 600 MHz Avance Bruker spectrometer with four rf channels and XYZ gradients. It also has a number of workstations for data acquisition and processing using many software packages and academic programs.

Abstracts of Current Research:

Dermaseptin Antimicrobial Peptides

Dermaseptins are a family of antimicrobial peptides, isolated from the skin of Phyllomedusa genus, South American tree frogs, which are considered to be an important component of the host defense system. Dermaseptin peptides show cytolytic activity in vitro against numerous microorganisms such as bacteria, protozoa, yeast and filamentous fungi.

NMR-derived structures of a large number of native dermaseptins and mutated, chemically modified and truncated analogues show a strong correlation between their degree of helicity and their biological activity. The helical structure of the peptides organizes the molecules such that they show amphipathic hydrophobicity and charge distribution that seem to be responsible for many aspects of their biological functions.

Dark Color Inducing Neurohormone Peptide

Dark color inducing neurohormone (DCIN) is responsible for the development of characteristic dark patterns in locusts as a result of densely populated living conditions. The color change is present when the locusts are in their gregarious phase at outbreaks ("locust plague").

The NMR-derived solution structure of this and similar peptides is under investigation in our laboratory and has shown a distinct electrostatic distribution that we expect to show the elements of the peptide that are essential for its biological activity.

Recent Publications (since 1995):

A. Zvi, I. Kustanovich, R. Levy, M. Eisenstein, Z. Matsushita, P. Richalet-Secordel, M. Regenmortel and J. Anglister, "Mapping of the Antigenic Determinant Recognized by an anti - gp120 HIV Neutralizing Antibody by Two-Dimensional NMR", Europen. J. Biochem. 229 (1), 178-87 (1995).

A. Zvi, I. Kustanovich, Y. Hayek, S. Matsushita and J. Anglister, "The Principal Neutralizing Determinant of HIV-1 Located in V3 of gp120 Forms a 12-Residue Loop by Internal Hydrophobic Interactions", FEBS Lett. 368 (2), 267-70 (1995).

I. Kustanovich and A. Zvi, "Epitope Mapping Antibody - Antigen Complexes by Nuclear Magnetic Resonance Spectroscopy", in "Methods in Molecular Biology", Vol. 66, p. 25-37 Ed. G. Morris, Humana Press Inc (1996).

V. Tugarinov, I. Kustanovich, N. Zilberberg, M. Gurevitz and J. Anglister, "Solution Structure of a Highly Insecticidal a-neurotoxin from the Scorpion Leiurus Quinquestriatus Hebraeus", Biochemistry 36 (9), 2414-24 (1997).

J. K. Ghosh, D. Shaaool, P. Guillaud, L. Ciceron, D. Mazier, I. Kustanovich, Y. Shai and A. Mor, "Selective Cytotoxicity of Dermaseptin S3 Towards Intraerythrocytic Plasmodium Falciparum and the Underlined Molecular Basis", J. Biol. Chem. 272 (50), 31609-16 (1997).

A. P. Hinck, M. A. Markus, S. Huang, S. Grzesiek, I. Kustanovich, D. E. Draper and D. A. Torchia, "The RNA Binding Domain of Ribosomal protein L11: Three Dimensional Structure of the RNA-bound Form of the Protein and its Interaction with 23S rRNA", J. Mol. Biol. 274 (1), 101-13 (1997).

J. Ghosh, I. Kustanovich, Y. Shai and A. Mor, "Selective Cytotoxicity of Dermaseptins", Proceeding book "First International Peptide Symposium", Ed. Y. Shimonishi "Peptide Science - Present and Future", p.725-7 (1998).

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