Topics in Spatial Econometrics - With Applications to House Prices

Topics in Spatial Econometrics — With Applications to House Prices Spatial effects in data occur when geographical closeness of observations influences the relation between the observations. When two points on a map are close to each other, the observed values on a variable at those points tend to be similar. The further away the two points are from each other, the less similar the observed values tend to be. Recent technical developments, geographical information systems (GIS) and global positioning systems (GPS) have brought about a renewed interest in spatial matters. For instance, it is possible to observe the exact location of an observation and combine it with other characteristics. Spatial econometrics integrates spatial aspects into econometric models and analysis. The thesis concentrates mainly on methodological issues, but the findings are illustrated by empirical studies on house price data. The thesis consists of an introductory chapter and four essays. The introductory chapter presents an overview of topics and problems in spatial econometrics. It discusses spatial effects, spatial weights matrices, especially k-nearest neighbours weights matrices, and various spatial econometric models, as well as estimation methods and inference. Further, the problem of omitted variables, a few computational and empirical aspects, the bootstrap procedure and the spatial J-test are presented. In addition, a discussion on hedonic house price models is included. In the first essay a comparison is made between spatial econometrics and time series analysis. By restricting the attention to unilateral spatial autoregressive processes, it is shown that a unilateral spatial autoregression, which enjoys similar properties as an autoregression with time series, can be defined. By an empirical study on house price data the second essay shows that it is possible to form coordinate-based, spatially autoregressive variables, which are at least to some extent able to replace the spatial structure in a spatial econometric model. In the third essay a strategy for specifying a k-nearest neighbours weights matrix by applying the spatial J-test is suggested, studied and demonstrated. In the final fourth essay the properties of the asymptotic spatial J-test are further examined. A simulation study shows that the spatial J-test can be used for distinguishing between general spatial models with different k-nearest neighbours weights matrices. A bootstrap spatial J-test is suggested to correct the size of the asymptotic test in small samples.

Eteva Järvenpään perhepalvelukeskuksen lyhytaikaishoidon suunnittelun kehittäminen

Objectives. The thesis objective was to analyze how person-centred planning is applied to develop short term care in interaction between the disabled children, their families and the workers of the family service centre of Eteva Järvenpää. The thesis contributes to developing the methods of person-centred planning. I applied theoretical frameworks of activity theory and developmental work research, family-based work framework and disability phenomenon. The research questions were: What development needs did the families of disabled children have for the services? How were viewpoints of disabled children, their families and Eteva workers noticed in person-centred planning in the interaction between the disabled children, their families and Eteva workers? What disturbances and development challenges emerged during the person-centred planning? Methods. I first analysed the local history of the disability sector and the short term care to analyse challenges arising from the local history. The actural research material consisted of interviews with four families, two person-centred planning discussions and two discussions where the person-centred planning was reflected by the families.I used interaction voice analysis as defined by the activity theory and developmental work research. From the recorded interviews and discussions I analysed scripts, disturbances, innovation attempts and innovations. From the discussions I analysed also the interaction types (cooperation, coordination and communication). Results and conclusions. As problems, the families considered the scarce resources and the inflexibility of services. The challenges of developing the short term care were how to transfer information from short term care to home, how to develop activities for the children and how to take into account the individual needs of the children in the short term care. Both from the local history analysis and from the family interviews arised the conflict between caring and fulfilling the individual needs. In person-centred planning, the voice of the child was either interpreted by other family members or guided by family members or workers. I modelled the progress of person-centred planning in a two-dimensional coordination. Person-centred planning should be deepened in cooperation between the child, the family and the workers in everyday situations at home and during the short term care. The challenge is to expand person-centred planning to become cross-organizational cooperation connecting the actors of the child s service network in everyday life. Avainsanat Nyckelord - Keywords short term care, activity theory and developmental work research, person-centred planning, disability

Subclinical autonomic neuropathy in alcoholics: Decreased heart rate variability

Cardiac autonomic neuropathy is known to occur in alcoholics but the extent of its subclinical form is not usually recognized, Heart Rate Variability (HRV) analysis can detect subclinical autonomic neuropathy. In this study the HRV parameters were compared in 20 neurologically asymptomatic alcoholics, 20 age-matched normals and 16 depressives. All were males, ECG was recorded in a quiet room for four minutes in supine position. Time and Frequency domain parameters of HRV were computed by a researcher blind to clinical details. Alcoholics had significantly smaller Coefficient of Variation of R-R intervals (CVR-R) on time domain analysis and smaller HF band (0.15-0.5 Hz) power on spectral analysis. The decreased Heart Rate Variability indicates cardiac autonomic dysfunction.

On Convergence of Differential Evolution Over a Class of Continuous Functions With Unique Global Optimum

Differential evolution (DE) is arguably one of the most powerful stochastic real-parameter optimization algorithms of current interest. Since its inception in the mid 1990s, DE has been finding many successful applications in real-world optimization problems from diverse domains of science and engineering. This paper takes a first significant step toward the convergence analysis of a canonical DE (DE/rand/1/bin) algorithm. It first deduces a time-recursive relationship for the probability density function (PDF) of the trial solutions, taking into consideration the DE-type mutation, crossover, and selection mechanisms. Then, by applying the concepts of Lyapunov stability theorems, it shows that as time approaches infinity, the PDF of the trial solutions concentrates narrowly around the global optimum of the objective function, assuming the shape of a Dirac delta distribution. Asymptotic convergence behavior of the population PDF is established by constructing a Lyapunov functional based on the PDF and showing that it monotonically decreases with time. The analysis is applicable to a class of continuous and real-valued objective functions that possesses a unique global optimum (but may have multiple local optima). Theoretical results have been substantiated with relevant computer simulations.

Northeast monsoon over India: variability and prediction

South peninsular India experiences a large portion of the annual rainfall during the northeast monsoon season (October to December). In this study, the facets of diurnal, intra-seasonal and inter-annual variability of the northeast monsoon rainfall (the NEMR) over India have been examined. The analysis of satellite derived hourly rainfall reveals that there are distinct features of diurnal variation over the land and oceans during the season. Over the land, rainfall peaks during the late afternoon/evening, while over the oceans an early morning peak is observed. The harmonic analysis of hourly data reveals that the amplitude and variance are the largest over south peninsular India. The NEMR also exhibits significant intra-seasonal variability on a 20-40 day time scale. Analysis also shows significant northward propagation of the maximum cloud zone from south of equator to the south peninsula during the season. The NEMR exhibits large inter-annual variability with the co-efficient of variation (CV) of 25%. The positive phases of ENSO and the Indian Ocean Dipole (IOD) are conducive for normal to above normal rainfall activity during the northeast monsoon. There are multi-decadal variations in the statistical relationship between ENSO and the NEMR. During the period 2001-2010 the statistical relationship between ENSO and the NEMR has significantly weakened. The analysis of seasonal rainfall hindcasts for the period 1960-2005 produced by the state-of-the-art coupled climate models, ENSEMBLES, reveals that the coupled models have very poor skill in predicting the inter-annual variability of the NEMR. This is mainly due to the inability of the ENSEMBLES models to simulate the positive relationship between ENSO and the NEMR correctly. Copyright (C) 2012 Royal Meteorological Society

Establishment of an in vitro transcription system for Peste des petits ruminant virus

Background: Peste-des-petits ruminants virus (PPRV) is a non segmented negative strand RNA virus of the genus Morbillivirus within Paramyxoviridae family. Negative strand RNA viruses are known to carry nucleocapsid (N) protein, phospho (P) protein and RNA polymerase (L protein) packaged within the virion which possess all activities required for transcription, post-transcriptional modification of mRNA and replication. In order to understand the mechanism of transcription and replication of the virus, an in vitro transcription reconstitution system is required. In the present work, an in vitro transcription system has been developed with ribonucleoprotein (RNP) complex purified from virus infected cells as well as partially purified recombinant polymerase (L-P) complex from insect cells along with N-RNA (genomic RNA encapsidated by N protein) template isolated from virus infected cells. Results: RNP complex isolated from virus infected cells and recombinant L-P complex purified from insect cells was used to reconstitute transcription on N-RNA template. The requirement for this transcription reconstitution has been defined. Transcription of viral genes in the in vitro system was confirmed by PCR amplification of cDNAs corresponding to individual transcripts using gene specific primers. In order to measure the relative expression level of viral transcripts, real time PCR analysis was carried out. qPCR analysis of the transcription products made in vitro showed a gradient of polarity of transcription from 3' end to 5' end of the genome similar to that exhibited by the virus in infected cells. Conclusion: This report describes for the first time, the development of an in vitro transcription reconstitution system for PPRV with RNP complex purified from infected cells and recombinant L-P complex expressed in insect cells. Both the complexes were able to synthesize all the mRNA species in vitro, exhibiting a gradient of polarity in transcription.

Auditory-motivated Gammatone wavelet transform

The ability of the continuous wavelet transform (CWT) to provide good time and frequency localization has made it a popular tool in time-frequency analysis of signals. Wavelets exhibit constant-Q property, which is also possessed by the basilar membrane filters in the peripheral auditory system. The basilar membrane filters or auditory filters are often modeled by a Gammatone function, which provides a good approximation to experimentally determined responses. The filterbank derived from these filters is referred to as a Gammatone filterbank. In general, wavelet analysis can be likened to a filterbank analysis and hence the interesting link between standard wavelet analysis and Gammatone filterbank. However, the Gammatone function does not exactly qualify as a wavelet because its time average is not zero. We show how bona fide wavelets can be constructed out of Gammatone functions. We analyze properties such as admissibility, time-bandwidth product, vanishing moments, which are particularly relevant in the context of wavelets. We also show how the proposed auditory wavelets are produced as the impulse response of a linear, shift-invariant system governed by a linear differential equation with constant coefficients. We propose analog circuit implementations of the proposed CWT. We also show how the Gammatone-derived wavelets can be used for singularity detection and time-frequency analysis of transient signals. (C) 2013 Elsevier B.V. All rights reserved.

An extended finite element model coupled with level set method for stable pitting corrosion

Mass balance between metal and electrolytic solution, separated by a moving interface, in stable pit growth results in a set of governing equations which are solved for concentration field and interface position (pit boundary evolution), which requires only three inputs, namely the solid metal concentration, saturation concentration of the dissolved metal ions and diffusion coefficient. A combined eXtended Finite Element Model (XFEM) and level set method is developed in this paper. The extended finite element model handles the jump discontinuity in the metal concentrations at the interface, by using discontinuous-derivative enrichment formulation for concentration discontinuity at the interface. This eliminates the requirement of using front conforming mesh and re-meshing after each time step as in conventional finite element method. A numerical technique known as level set method tracks the position of the moving interface and updates it over time. Numerical analysis for pitting corrosion of stainless steel 304 is presented. The above proposed method is validated by comparing the numerical results with experimental results, exact solutions and some other approximate solutions.

Part I: High-Voltage MOS Device Design for Improved Static and RF Performance

In this paper, for the first time, the key design parameters of a shallow trench isolation-based drain-extended MOS transistor are discussed for RF power applications in advanced CMOS technologies. The tradeoff between various dc and RF figures of merit (FoMs) is carefully studied using well-calibrated TCAD simulations. This detailed physical insight is used to optimize the dc and RF behavior, and our work also provides a design window for the improvement of dc as well as RF FoMs, without affecting the breakdown voltage. An improvement of 50% in R-ON and 45% in RF gain is achieved at 1 GHz. Large-signal time-domain analysis is done to explore the output power capability of the device.

Tetraphenylethene-Based Conjugated Fluoranthene: A Potential Fluorescent Probe for Detection of Nitroaromatic Compounds

This study reports the synthesis and photophysical properties of a star-shaped, novel, fluoranthene-tetraphenylethene (TFPE) conjugated luminogen, which exhibits aggregation-induced blue-shifted emission (AIBSE). The bulky fluoranthene units at the periphery prevent intramolecular rotation (IMR) of phenyl rings and induces a blueshift with enhanced emission. The AIBSE phenomenon was investigated by solvatochromic and temperature-dependent emission studies. Nanoaggregates of TFPE, formed by varying the water/THF ratio, were investigated by SEM and TEM and correlated with optical properties. The TFPE conjugate was found to be a promising fluorescent probe towards the detection of nitroaromatic compounds (NACs), especially for 2,4,6-trinitrophenol (PA) with high sensitivity and a high Stern-Volmer quenching constant. The study reveals that nanoaggregates of TFPE formed at 30 and 70% water in THF showed unprecedented sensitivity with detection limits of 0.8 and 0.5ppb, respectively. The nanoaggregates formed at water fractions of 30 and 70% exhibit high Stern-Volmer constants (K-sv=79998 and 51120m(-1), respectively) towards PA. Fluorescence quenching is ascribed to photoinduced electron transfer between TFPE and NACs with a static quenching mechanism. Test strips coated with TFPE luminogen demonstrate fast and ultra-low-level detection of PA for real-time field analysis.

Proceedings of the 1998 Science Board Symposium on The Impacts of the 1997/98 El Niño Event on the North Pacific Ocean and Its Marginal Seas

Preface [pdf, 0.01 Mb] James J. O'Brien The big picture - The ENSO of 1997-98 [pdf, 0.01 Mb] James E. Overland, Nicholas A. Bond & Jennifer Miletta Adams Atmospheric anomalies in 1997: Links to ENSO? [pdf, 0.54 Mb] Vladimir I. Ponomarev, Olga Trusenkova, Serge Trousenkov, Dmitry Kaplunenko, Elena Ustinova & Antonina Polyakova The ENSO signal in the northwest Pacific [pdf, 0.47 Mb] Robert L. Smith, A. Huyer, P.M. Kosro & J.A. Barth Observations of El Niño off Oregon: July 1997 to present (October 1998) [pdf, 1.31 Mb] Patrica A. Wheeler & Jon Hill Biological effects of the 1997-1998 El Niño event off Oregon: Nutrient and chlorophyll distributions [pdf, 1.13 Mb] William T. Peterson Hydrography and zooplankton off the central Oregon coast during the 1997-1998 El Niño event [pdf, 0.26 Mb] William Crawford, Josef Cherniawsky, Michael Foreman & Peter Chandler El Niño sea level signal along the west coast of Canada [pdf, 1.25 Mb] Howard J. Freeland & Rick Thomson The El Niño signal along the west coast of Canada - temperature, salinity and velocity [pdf, 0.49 Mb] Frank A. Whitney, David L. Mackas, David W. Welch & Marie Robert Impact of the 1990s El Niños on nutrient supply and productivity of Gulf of Alaska waters [pdf, 0.06 Mb] Craig McNeil, David Farmer & Mark Trevorrow Dissolved gas measurements at Stn. P4 during the 97-98 El Niño [pdf, 0.13 Mb] Kristen L.D. Milligan, Colin D. Levings & Robert E. DeWreede Data compilation and preliminary time series analysis of abundance of a dominant intertidal kelp species in relation to the 1997/1998 El Niño event [pdf, 0.05 Mb] S.M. McKinnell, C.C. Wood, M. Lapointe, J.C. Woodey, K.E. Kostow, J. Nelson & K.D. Hyatt Reviewing the evidence that adult sockeye salmon strayed from the Fraser River and spawned in other rivers in 1997 [pdf,0.03 Mb] G.A. McFarlane & R.J. Beamish Sardines return to British Columbia waters [pdf, 0.34 Mb] Ken H. Morgan Impact of the 1997/98 El Niño on seabirds of the northeast Pacific [pdf, 0.06 Mb] Thomas C. Royer & Thomas Weingartner Coastal hydrographic responses in the northern Gulf of Alaska to the 1997-98 ENSO event [pdf, 0.76 Mb] John F. Piatt, Gary Drew, Thomas Van Pelt, Alisa Abookire, April Nielsen, Mike Shultz & Alexander Kitaysky Biological effects of the 1997/98 ENSO in Cook Inlet, Alaska [pdf, 0.22 Mb] H.J. Niebauer The 1997-98 El Niño in the Bering Sea as compared with previous ENSO events and the "regime shift" of the late 1970s [pdf, 0.10 Mb] A.S. Krovnin, G.P. Nanyushin, M.Yu. Kruzhalov, G.V. Khen, M.A. Bogdanov, E.I. Ustinova, V.V. Maslennikov, A.M. Orlov, B.N. Kotenev, V.V. Bulanov & G.P. Muriy The state of the Far East seas during the 1997/98 El Niño event [pdf, 0.15 Mb] Stacy Smith & Susan Henrichs Phytoplankton collected by a time-series sediment trap deployed in the southeast Bering Sea during 1997 [pdf, 0.21 Mb] Cynthia T. Tynan Redistributions of cetaceans in the southeast Bering Sea relative to anomalous oceanographic conditions during the 1997 El Niño [pdf, 0.02 Mb] Akihiko Yatsu, Junta Mori, Hiroyuki Tanaka, Tomowo Watanabe, Kazuya Nagasawa, Yikimasa Ishida, Toshimi Meguro, Yoshihiko Kamei & Yasunori Sakurai Stock abundance and size compositions of the neon flying squid in the central North Pacific Ocean during 1979-1998 [pdf, 0.11 Mb] O.B. Feschenko A new point of view concerning the El Niño mechanism [pdf, 0.01 Mb] Nathan Mantua 97/98 Ocean climate variability in the northeast Pacific: How much blame does El Niño deserve? [pdf, 0.01 Mb] Vadim P. Pavlychev Sharp changes of hydrometeorological conditions in the northwestern Pacific during the 1997/1998 El Niño event [pdf, 0.01 Mb] Jingyi Wang Predictability and forecast verification of El Niño events [pdf, 0.01 Mb] (Document contains 110 pages)

Streamflow response of a small forested catchment on different timescales

The hydrological response of a catchment to rainfall on different timescales is result of a complex system involving a range of physical processes which may operate simultaneously and have different spatial and temporal influences. This paper presents the analysis of streamflow response of a small humid-temperate catchment (Aixola, 4.8 km(2)) in the Basque Country on different timescales and discusses the role of the controlling factors. Firstly, daily time series analysis was used to establish a hypothesis on the general functioning of the catchment through the relationship between precipitation and discharge on an annual and multiannual scale (2003-2008). Second, rainfall-runoff relationships and relationships among several hydrological variables, including catchment antecedent conditions, were explored at the event scale (222 events) to check and improve the hypothesis. Finally, the evolution of electrical conductivity (EC) during some of the monitored storm events (28 events) was examined to identify the time origin of waters. Quick response of the catchment to almost all the rainfall events as well as a considerable regulation capacity was deduced from the correlation and spectral analyses. These results agree with runoff event scale data analysis; however, the event analysis revealed the non-linearity of the system, as antecedent conditions play a significant role in this catchment. Further, analysis at the event scale made possible to clarify factors controlling (precipitation, precipitation intensity and initial discharge) the different aspects of the runoff response (runoff coefficient and discharge increase) for this catchment. Finally, the evolution of EC of the waters enabled the time origin (event or pre-event waters) of the quickflow to be established; specifically, the conductivity showed that pre-event waters usually represent a high percentage of the total discharge during runoff peaks. The importance of soil waters in the catchment is being studied more deeply.

Distribution of skipjack in the Pacific Ocean, based on records of incidental catches of the Japanese longline tuna fishery

ENGLISH: All available longline data on skipjack captured in the Pacific Ocean by Japanese research vessels (1949-1965) and from incidental skipjack catches by Japanese commercial vessels (1956-1964) were analyzed. As skipjack are not specifically sought by longline vessels, the data are limited. Considering this it was found that: longline gear captures skipjack of wider size-range and is more selective for larger skipjack than conventional fishing methods, i.e. pole-and-line and purse-seine; skipjack are widely and almost continuously distributed across the Pacific; throughout the year average hook-rates are greater in the southeastern Pacific than in the northwestern Pacific; areas of high hook-rate shift south during the second and third quarters and north during the first and fourth quarters; in the western Pacific the north-south range of the catch distribution was greatest in the first and fourth quarters; skipjack hook-rates are relatively high in the northwestern Pacific east of Japan only during the first and fourth quarters; the highest hook-rates were recorded in extensive areas along the equator (from lO°N to 20°8 between approximately 155°W-100°W); generally more skipjack were captured by research longline gear in water temperature ranges approaching both the upper and lower temperature limits of skipjack distribution (18-21C and 26-28C), than is the case in surface skipjack fisheries; tentative comparisons of longline skipjack catch distributions with Pacific current systems, suggests low skipjack abundance in both North Pacific Central and North Pacific Equatorial water; the sex ratio was 95 males : 63 females in a small sample of skipjack examined; longlines capture skipjack of three, and possibly more, age groups; in skipjack size-composition samples studied, the smaller modal group (65 cm) observed in January-March in the northwestern Pacific (1600E-180oE and 20oN-45°N) corresponds in size to the larger modal group appearing in the late-summer surface fishery off the Izu-Bonin Islands southeast of Japan, and also compares in modal size to the skipjack taken in the Hawaiian fishery in spring time; the analysis of skipjack catches by hook position on the longline and by death-rate studies, indicates that part of the catch is made while the gear is in motion near the surface, and a lesser part of the catch is made when the gear is stabilized at a depth of 70 to 140 m. A brief discussion is given, in the light of new information presented, on several hypotheses by other authors concerning the population structure and migration of skipjack in the Pacific Ocean. SPANISH: Se analizaron todos los datos disponibles de la pesca con palangre de barriletes capturados en el Océano Pacífico por barcos japoneses de investigación (1949-1965) y por las capturas incidentales de los barcos comerciales japoneses (1956-1964). Como los barcos palangreros específicamente, no persiguen al barrilete, los datos son limitados. Considerando ésto, se encontró: que el arte palangrero obtiene barriletes con una distribución más amplia de tallas, y es más selectivo en cuanto a los barriletes de mayor talla, que los métodos convencionales de pesca, Le. cañas de pescar y redes de cerco; el barrilete se encuentra amplia y casi continuamente distribuido a través del Pacífico; en todo el año, las tasas promedio de captura por anzuelo son superiores en el Pacífico sudoriental que las del Pacífico noroeste; las áreas con una tasa alta de captura por anzuelo, se cambian hacia el sur durante los trimestres segundo y tercero, y durante los trimestres primero y cuarto hacia el norte; en el Pacífico occidental la amplitud de la distribución de captura norte-sur, fue superior en los trimestres primero y cuarto; las tasas de captura por anzuelo de barrilete, son relativamente altas en el Pacífico noroeste al este del Japón, únicamente durante los trimestres primero y cuarto; las tasas de captura por anzuelo más altas fueron registradas en extensas áreas a lo largo del ecuador (desde los 10°N hasta los 20°S, aproximadamente entre los 155°W-100°W) ; generalmente las artes palangreras de investigación capturaron más barrilete en aguas en las que la temperatura se aproximaba a los límites más altos o bajos de la temperatura en la distribución del barrilete (18-21 C y 26-28 C), que en el caso de la pesca superficial de barrilete; las comparaciones tentativas de la captura de barrilete con palangre, con el sistema de las corrientes del Pacífico, sugieren una abundancia inferior de barrilete tanto en las aguas del Pacífico central del norte como en las del Pacífico ecuatorial del norte; la proporcíon sexual examinada en una pequeña muestra de barriletes, fue de 95 machos y 63 hembras; los palangreros capturan barriletes de tres grupos de edad y posiblemente de más; en las muestras estudiadas de la composición de las tallas de barrilete, el grupo modal más pequeño (65 cm), observado en enero-marzo en el Pacífico noroeste (160 0E-180° y 20 oN-45°N), corresponde en talla al grupo modal más grande que aparece en la pesca de superficie a fines del verano frente a las Islas Izu-Bonín al sudeste del Japón, y se compara también con la talla modal del barrilete obtenido en la pesca hawaiana en la época de primavera; el análisis de las capturas de barrilete por medio del estudio de la posición de los anzuelos en el palangre y por la tasa de mortalidad, indica que parte de la captura se efectúa cuando el equipo está en movimiento cerca a la superficie y una parte inferior de la captura se realiza, cuando las artes se estabilizan a una profundidad de 70 a 140 m. Se ofrece una breve discusión sobre varias hipótesis de otros autores, en vista de la nueva información presentada referente a la estructura poblacional y a la migración del barrilete en el Océano Pacífico. (PDF contains 100 pages.)

Two-dimensional Vortex-induced Vibrations of Submerged Floating Tunnel Tethers

A vortex-induced vibration (VIV) model is presented for predicting the nonlinear dynamic response of submerged floating tunnel (SFT) tethers which are subjected to wave, current and tunnel oscillatory displacements at their upper end in horizontal and vertical directions. A nonlinear fluid force formula is introduced in this model, and the effect of the nonlinearity of tether is investigated. First, the tunnel is stationary and the tether vibrates due to the vortices shedding. The calculated results show that the cross-flow amplitude of VIV decreases compared with the linear model. However the in-line amplitude of VIV increases. Next, the periodical oscillation of tunnel is considered. The oscillation caused by wave forces plays the roles of parametric exciter and forcing exciter to the VIV of tether. The time history of displacement of the tether mid-span is obtained by the proposed model. It is shown that the in-line amplitude increases obviously and the corresponding frequency is changed. The cross-flow amplitude exhibits a periodic behavior.

Structural requirements for catalysis: studies on RTEM-1 β-lactamase

β-lactamases are a group of enzymes that confer resistance to penam and cephem antibiotics by hydrolysis of the β-lactam ring, thereby inactivating the antibiotic. Crystallographic and computer modeling studies of RTEM-1 β-lactamase have indicated that Asp 132, a strictly conserved residue among the class A β-lactamases, appears to be involved in substrate binding, catalysis, or both. To study the contribution of residue 132 to β-lactamase function, site saturation mutagenesis was used to generate mutants coding for all 20 amino acids at position 132. Phenotypic screening of all mutants indicated that position 132 is very sensitive to amino acid changes, with only N132C, N132D, N132E, and N132Q showing any appreciable activity. Kinetic analysis of three of these mutants showed increases in K_M, along with substantial decreases in k_(cat). Efforts to trap a stable acyl-enzyme intermediate were unsuccessfuL These results indicate that residue 132 is involved in substrate binding, as well as catalysis, and supports the involvement of this residue in acylation as suggested by Strynadka et al.

Crystallographic and computer modeling studies of RTEM-1 β-lactamase have indicated that Lys 73 and Glu 166, two strictly conserved residues among the class A β-lactamases, appear to be involved in substrate binding, catalysis, or both. To study the contribution of these residues to β-lactamase function, site saturation mutagenesis was used to generate mutants coding for all 20 amino acids at positions 73 and 166. Then all 400 possible combinations of mutants were created by combinatorial mutagenesis. The colonies harboring the mutants were screened for growth in the presence of ampicillin. The competent colonys' DNA were sequenced, and kinetic parameters investigated. It was found that lysine is essential at position 73, and that position 166 only tolerated fairly conservative changes (Aspartic acid, Histidine, and Tyrosine). These functional mutants exhibited decreased kcat's, but K_M was close to wild-type levels. The results of the combinatorial mutagenesis experiments indicate that Lysis absolutely required for activity at position 73; no mutation at residue 166 can compensate for loss of the long side chain amine. The active mutants found--K73K/E166D, K73KIE166H, and K73KIE166Y were studied by kinetic analysis. These results reaffirmed the function of residue 166 as important in catalysis, specifically deacylation.

The identity of the residue responsible for enhancing the active site serine (Ser 70) in RTEM-1 β-lactamase has been disputed for some time. Recently, analysis of a crystal structure of RTEM-1 β-lactamase with covalently bound intermediate was published, and it was suggested that Lys 73, a strictly conserved residue among the class A β-lactamases, was acting as a general base, activating Ser 70. For this to be possible, the pK_a of Lys 73 would have to be depressed significantly. In an attempt to assay the pK_a of Lys 73, the mutation K73C was made. This mutant protein can be reacted with 2-bromoethylamine, and activity is restored to near wild type levels. ^(15)N-2-bromoethylamine hydrobromide and ^(13)C-2-bromoethylamine hydrobromide were synthesized. Reacting these compounds with the K73C mutant gives stable isotopic enrichment at residue 73 in the form of aminoethylcysteine, a lysine homologue. The pK_a of an amine can be determined by NMR titration, following the change in chemical shift of either the ^(15)N-amine nuclei or adjacent Be nuclei as pH is changed. Unfortunately, low protein solubility, along with probable label scrambling in the Be experiment, did not permit direct observation of either the ^(15)N or ^(13)C signals. Indirect detection experiments were used to observe the protons bonded directly to the ^(13)C atoms. Two NMR signals were seen, and their chemical shift change with pH variation was noted. The peak which was determined to correspond to the aminoethylcysteine residue shifted from 3.2 ppm down to 2.8 ppm over a pH range of 6.6 to 12.5. The pK_a of the amine at position 73 was determined to be ~10. This indicates that residue 73 does not function as a general base in the acylation step of the reaction. However the experimental measurement takes place in the absence of substrate. Since the enzyme undergoes conformational changes upon substrate binding, the measured pK_a of the free enzyme may not correspond to the pK_a of the enzyme substrate complex.