Engineering cofactor specificity of methyltransferases

Filarete`s Libro architettonico, written in Milan around 1460/65, has so far been almost exclusively studied by art historians, who have always treated the text as if it were merely a treatise on architecture. Anything that could not be assigned to the later tradition of the genre of the architectural treatise initiated by Alberti (or that could not be linked to actual buildings) has been prematurely dismissed as `abstruse`, `fantastic` or `romantic`, or at best as an insignificant digression used only to keep the reader entertained. The research project is concerned with previously little-noted or overlooked passages. Through a precise reading and an interdisciplinary study of source texts going well beyond the usual art-historical reference points including astronomical/astrological textbooks, the hermetic tradition, manuals of magic, travel literature, and other genres , it is intended to show that Filarete`s Libro architettonico is an outstanding document that crystallizes its time, in the transition between the Middle Ages and the early modern period. A longer stay at the Kunsthistorisches Institut Florenz (Max-Planck-Institute) would enable the applicant (1.) to systematically search the rich holdings of the Florentine archives for information about Filarete (Archivio di Stato di Firenze, registries of births, the fraternity files of the sacred institutions), whose origins, year of birth and early career in Florence before he moved to Rome in 1433 (or earlier?) and eventually to Milan in 1451 are completely obscure; (2.) to investigate Filarete`s position within the Humanist milieu in Florence, to which substantial parts of his Libro architettonico refer back. The project of comprehensively reconstructing these original influences or web of contexts concerns Filarete`s contacts with Giovanni di Cosimo and Piero de` Medici and members of their Humanist circle such as Niccolò Niccoli and Ciriaco d`Ancona or Benedetto Dei, or the Sienese Beltramo Mignanelli. Secondly, it is concerned with the range of manuscripts available to Filarete in the early 15th century, as preserved, above all, in the Biblioteca Laurenziana, the Biblioteca Riccardiana, and the Biblioteca Nazionale Centrale. This aspect is concerned in particular with reconstructing the range of Arabic manuscripts available to Filarete. The project is intended to lead to an extensive monograph that will provide a synthetic presentation of the numerous aspects of the Libro architettonico and its author in the context of the cultural history of the early Renaissance between Milan and Florence, the Middle Ages and the Renaissance. Another aim of the project is also to obtain authorization ("Venia Legendi") from the University of Vienna to teach art history (on completion of my postdoctoral Habilitation degree).

In the course of the project Engineering Cofactor Specificity of Methyltransferases a novel experimental approach for describing selection in directed evolution, which is performed in the laboratory, was found. Directed evolution aims to isolate new protein variants in a very fast manner compared to natural evolution. One important difference is that directed evolution only focuses on one protein under isolated conditions, which enables scientists to isolate new variants within a short period of time. Research so far has focused on the output of the directed evolution experiment, while the molecular level of the selection process has never been investigated. We made use of the emergent tool of the so-called Next Generation Sequencing technology, which makes it possible to analyze single DNA molecules within a pool of DNA. By this means we could clearly monitor the impact of the selective pressure, in this particular case the concentration of the proteins interaction partner, on the survival of certain DNA species. A very interesting finding was that a low concentration of the interaction partner results in a strong selection of the wild-type species. In other words, if there is a low availability of food, only the most-adapted species can survive. In contrast to that almost no selection occurs at a high concentration of the interaction partner, indicating the survival of a broad range of species in case of a surplus of food. However it was found, if new variants with new activities are to be isolated, a surplus of the non-natural interaction partner needs to be present. Potentially a new sort of food needs to be available in huge amounts in order to feed new species. Theoretically a bigger population could also be advantageous. Our approach made it possible to study the mechanism of evolutionary selection on a single molecule level, which is novel to the scientific community. Apart from the evolutionary studies, rational protein design was used in order to identify areas of a protein that are important for the binding to the interaction partner. The analyses of several protein variants and a number of different interaction partners provided information about binding strength and important areas of both the protein and the interaction partner. These findings are of particular interest for the development of new methodologies for protein-interaction partner studies and for the development of new biological tools, which are demanded in the chemical and pharmaceutical industry.