Active Site Structures of CYP11A1 in the Presence of Its Physiological Substrates and Alterations upon Binding of Adrenodoxin

The rate-limiting step in the steroid synthesis pathway is catalyzed by CYP11A1 through three sequential reactions. The first two steps involve hydroxylations at positions 22 and 20, generating 20(R),22(R)-dihydroxycholesterol (20R,22R-DiOHCH), with the third stage leading to a C20–C22 bond cleavage, forming pregnenolone. This work provides detailed information about the active site structure of CYP11A1 in the resting state and substrate-bound ferric forms as well as the CO-ligated adducts. In addition, high-quality resonance Raman spectra are reported for the dioxygen complexes, providing new insight into the status of Fe–O–O fragments encountered during the enzymatic cycle. Results show that the three natural substrates of CYP11A1 have quite different effects on the active site structure, including variations of spin state populations, reorientations of heme peripheral groups, and, most importantly, substrate-mediated distortions of Fe–CO and Fe–O2 fragments, as revealed by telltale shifts of the observed vibrational modes. Specifically, the vibrational mode patterns observed for the Fe–O–O fragments with the first and third substrates are consistent with H-bonding interactions with the terminal oxygen, a structural feature that tends to promote O–O bond cleavage to form the Compound I intermediate. Furthermore, such spectral data are acquired for complexes with the natural redox partner, adrenodoxin (Adx), revealing protein–protein-induced active site structural perturbations. While this work shows that Adx has an only weak effect on ferric and ferrous CO states, it has a relatively stronger impact on the Fe–O–O fragments of the functionally relevant oxy complexes

Lincoln Pilgrimage and Celebration: Celebration of Lincoln\u27s Birthday at Lincoln Memorial University, Harrogate, Tennessee, February 10-12, 1917

Reprinted from the Lincoln Herald, October, 1950, vol. 52, no. 3; Cover and caption title. 1000 copies. The story of his celebration ... re-told. --Page 1. The celebration was also the 20th anniversary of the chartering of the university. Includes a cartoon by E.E. Burtt, of the Knoxville (Tenn.) Journal & Tribune, depicting Lincoln\u27s spirit hovering over the university (p. 10); also includes Extracts of some of the more important tributes (p. 12-15).https://scholarsjunction.msstate.edu/fvw-pamphlets/1561/thumbnail.jp

Ultrasonic Synthetic-Aperture Holographic Imaging†

Ultrasonic reflection imaging has become an important tool in NDE [1,2,3]. The lateral resolution of such images is limited by the aperture size of the transducer, while the depth resolution is limited by the pulse length. In addition, for a given aperture diameter, the lateral resolution degrades with depth as given by the Rayleigh criterion. In this paper we discuss a method to increase the effective aperture through aperture synthesis. Waveform data, collected from a scanned transducer focused near the surface of the sample, is coherently processed to yield a synthesized aperture which can be focused to any depth with constant resolution. The synthetic aperture method allows efficient volume inspection by trading off scan time with processing time, the latter of which is constantly decreasing with increasing computing power

Rocket Ignition Demonstrations Using Silane

Rocket ignition demonstration tests using silane were performed at the Penn State Combustion Research Laboratory. A heat sink combustor with one injection element was used with gaseous propellants. Mixtures of silane and hydrogen were used as fuel, and oxygen was used as oxidizer. Reliable ignition was demonstrated using fuel lead and and a swirl injection element

Bacteria Source Tracking to Support Watershed Planning, Little Sac River, Southwest Missouri

The Little Sac Watershed in Greene and Polk Counties of southwest Missouri was placed on the 303d list for bacteria impairment in 1998 (WCO 2016). In 2006, a Total Maximum Daily Load (TMDL) was developed for the watershed to address bacteria impairments within the Little Sac River and an initial watershed management plan was finalized in 2010 (Baffaut 2006, WCO 2009). The Watershed Committee of the Ozarks (WCO) is presently updating that plan with the most recent information on bacteria within the watershed. As part of that process, the WCO has contracted the Ozarks Environmental and Water Resources Institute (OEWRI) at Missouri State University (MSU) to complete a bacteria source tracking study within the watershed to identify potential bacteria pollution sources. The purpose of this study is to collect water samples throughout the watershed and evaluate bacteria DNA using real-time PCR for specific marker genes that can help identify specific bacteria sources from different locations in the Little Sac River watershed

Exploration of factors driving incorporation of unnatural dNTPS into DNA by Klenow fragment (DNA polymerase I) and DNA polymerase α

In order to further understand how DNA polymerases discriminate against incorrect dNTPs, we synthesized two sets of dNTP analogues and tested them as substrates for DNA polymerase α (pol α) and Klenow fragment (exo(−)) of DNA polymerase I (Escherichia coli). One set of analogues was designed to test the importance of the electronic nature of the base. The bases consisted of a benzimidazole ring with one or two exocyclic substituent(s) that are either electron-donating (methyl and methoxy) or electron-withdrawing (trifluoromethyl and dinitro). Both pol α and Klenow fragment exhibit a remarkable inability to discriminate against these analogues as compared to their ability to discriminate against incorrect natural dNTPs. Neither polymerase shows any distinct electronic or steric preferences for analogue incorporation. The other set of analogues, designed to examine the importance of hydrophobicity in dNTP incorporation, consists of a set of four regioisomers of trifluoromethyl benzimidazole. Whereas pol α and Klenow fragment exhibited minimal discrimination against the 5- and 6-regioisomers, they discriminated much more effectively against the 4- and 7-regioisomers. Since all four of these analogues will have similar hydrophobicity and stacking ability, these data indicate that hydrophobicity and stacking ability alone cannot account for the inability of pol α and Klenow fragment to discriminate against unnatural bases. After incorporation, however, both sets of analogues were not efficiently elongated. These results suggest that factors other than hydrophobicity, sterics and electronics govern the incorporation of dNTPs into DNA by pol α and Klenow fragment

Design and calibration of percolation samplers for measuring polyacrylamide-amended furrow-irrigation effects on drainage water quality

Amending irrigation furrow inflows with polyacrylamide (PAM) at low concentrations (10 mg L -1 ) reduces irrigation-induced erosion by 94% and increases infiltration by 15%, relative to untreated furrows. We hypothesized that PAM erosion-control technology would allow irrigation managers to increase furrow inflows to speed furrow-stream advance, produce a more uniform water distribution down field, and reduce the leaching hazard at the upper end (due to reduced infiltration opportunity time and/or shorter sets). We developed, tested, and installed instruments in a furrow irrigated Portneuf silt loam (Co-Si, mixed, mesic, Durixerollic Calciorthids with 1.6% slope) to investigate this premise. Soils were instrumented with repeating pulse multivibrator (CS-615) soil water sensors, thermocouples, tensiometers, and percolation soil water samplers at upper and lower ends of the furrows. Percolation samplers consisted of a 23-cm-deep, 20-cm-dia. stainless-steel beaker with a 1 7-cm-long, 4-cm-dia., 0.5 bar air-entry ceramic cup imbedded in a 5-cm-deep silica flour layer, slurried into the beaker bottom. Water was collected under suction (~ 1.4x ambient) via teflon tubes. Percolation sampler design and testing, field installation, and study experimental design are discussed