Olefin cyclopropanation and carbon-hydrogen amination via carbene and nitrene transfers catalyzed by engineered cytochrome P450 enzymes

Synthetic biology promises to transform organic synthesis by enabling artificial catalysis in living cells. I start by reviewing the state of the art in this young field and recognizing that new approaches are required for designing enzymes that catalyze nonnatural reactions, in order to expand the scope of biocatalytic transformations. Carbene and nitrene transfers to C=C and C-H bonds are reactions of tremendous synthetic utility that lack biological counterparts. I show that various heme proteins, including cytochrome P450BM3, will catalyze promiscuous levels of olefin cyclopropanation when provided with the appropriate synthetic reagents (e.g., diazoesters and styrene). Only a few amino acid substitutions are required to install synthetically useful levels of stereoselective cyclopropanation activity in P450BM3. Understanding that the ferrous-heme is the active species for catalysis and that the artificial reagents are unable to induce a spin-shift-dependent increase in the redox potential of the ferric P450, I design a high-potential serine-heme ligated P450 (P411) that can efficiently catalyze cyclopropanation using NAD(P)H. Intact E. coli whole-cells expressing P411 are highly efficient asymmetric catalysts for olefin cyclopropanation. I also show that engineered P450s can catalyze intramolecular amination of benzylic C-H bonds from arylsulfonyl azides. Finally, I review other examples of where synthetic reagents have been used to drive the evolution of novel enzymatic activity in the environment and in the laboratory. I invoke preadaptation to explain these observations and propose that other man-invented reactions may also be transferrable to natural enzymes by using a mechanism-based approach for choosing the enzymes and the reagents. Overall, this work shows that existing enzymes can be readily adapted for catalysis of synthetically important reactions not previously observed in nature.

New technologies driving decade-bandwidth radio astronomy : quad-ridged flared horn and compound-semiconductor LNAs

Among the branches of astronomy, radio astronomy is unique in that it spans the largest portion of the electromagnetic spectrum, e.g., from about 10 MHz to 300 GHz. On the other hand, due to scientific priorities as well as technological limitations, radio astronomy receivers have traditionally covered only about an octave bandwidth. This approach of "one specialized receiver for one primary science goal" is, however, not only becoming too expensive for next-generation radio telescopes comprising thousands of small antennas, but also is inadequate to answer some of the scientific questions of today which require simultaneous coverage of very large bandwidths.

This thesis presents significant improvements on the state of the art of two key receiver components in pursuit of decade-bandwidth radio astronomy: 1) reflector feed antennas; 2) low-noise amplifiers on compound-semiconductor technologies. The first part of this thesis introduces the quadruple-ridged flared horn, a flexible, dual linear-polarization reflector feed antenna that achieves 5:1-7:1 frequency bandwidths while maintaining near-constant beamwidth. The horn is unique in that it is the only wideband feed antenna suitable for radio astronomy that: 1) can be designed to have nominal 10 dB beamwidth between 30 and 150 degrees; 2) requires one single-ended 50 Ohm low-noise amplifier per polarization. Design, analysis, and measurements of several quad-ridged horns are presented to demonstrate its feasibility and flexibility.

The second part of the thesis focuses on modeling and measurements of discrete high-electron mobility transistors (HEMTs) and their applications in wideband, extremely low-noise amplifiers. The transistors and microwave monolithic integrated circuit low-noise amplifiers described herein have been fabricated on two state-of-the-art HEMT processes: 1) 35 nm indium phosphide; 2) 70 nm gallium arsenide. DC and microwave performance of transistors from both processes at room and cryogenic temperatures are included, as well as first-reported measurements of detailed noise characterization of the sub-micron HEMTs at both temperatures. Design and measurements of two low-noise amplifiers covering 1--20 and 8—50 GHz fabricated on both processes are also provided, which show that the 1--20 GHz amplifier improves the state of the art in cryogenic noise and bandwidth, while the 8--50 GHz amplifier achieves noise performance only slightly worse than the best published results but does so with nearly a decade bandwidth.