Fis is a nucleoid-associated protein that interacts with poorly related DNA sequences with a high degree of specificity. A difference of more than 3 orders of magnitude in apparent K d values was observed between specific (K d , ϳ1 to 4 nM) and nonspecific (K d , ϳ4 M) DNA binding. To examine the contributions of Fis residues to the high-affinity binding at different DNA sequences, 13 alanine substitutions were generated in or near the Fis helix-turn-helix DNA binding motif, and the resulting proteins were purified. In vitro binding assays at three different Fis sites (fis P II, hin distal, and attR) revealed that R85, T87, R89, K90, and K91 played major roles in high-affinity DNA binding and that R85, T87, and K90 were consistently vital for binding to all three sites. Other residues made variable contributions to binding, depending on the binding site. N84 was required only for binding to the attR Fis site, and the role of R89 was dramatically altered by the attR DNA flanking sequence. The effects of Fis mutations on fis P II or hin distal site binding in vitro generally correlated with their abilities to mediate fis P repression or DNA inversion in vivo, demonstrating that the in vitro DNAbinding effects are relevant in vivo. The results suggest that while Fis is able to recognize a minimal common set of DNA sequence determinants at different binding sites, it is also equipped with a number of residues that contribute to the binding strength, some of which play variable roles.Fis (factor for inversion stimulation) is the most abundant nucleoid-associated protein during the logarithmic growth phase in rapidly growing Escherichia coli cells (3, 6). However, during mid-to late-logarithmic growth, the intracellular levels of Fis decrease over 500-fold and become nearly imperceptible during the stationary phase. The impact of Fis on cell physiology is widespread. As a nucleoid-associated protein, it is able to interact with a large number of DNA sites to alter DNA topology (66,67). Numerous genes are subject to positive or negative regulation by Fis directly or indirectly (6,10,19,22,30,49,56,61,(73)(74)(75). In the case of the promoters rrnB P1 and proP P2, Fis binding to sites centered at positions Ϫ71 and Ϫ41, respectively, directly stimulates transcription by contacting the C-terminal domain of the RNA polymerase ␣ subunit (␣-CTD) (9, 43). In the case of the fis promoter (fis P), Fis binding to sites I and II, centered at positions ϩ25 and Ϫ44, respectively, negatively autoregulates the fis operon by hindering RNA polymerase binding (6, 50, 58). As its name suggests, Fis also stimulates site-specific DNA inversion involving the Hin, Gin, and Cin family of recombinases (24,32,35). During Hin-mediated DNA inversion, Fis binds to two high-affinity DNA sites (hin-proximal and -distal sites) in a 60-bp recombinational enhancer region and interacts with two DNA-bound Hin recombinases to form a nucleoprotein complex intermediate required for efficient DNA strand cleavage and inversion (25,33). Bacteriophage DNA excis...

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The paper presents the theoretical development of the equations of motion for the flexural vibrations of a shallow spherical shell of orthotropic material. For symmetrical vibrations, the motion is described by two ordinary sixth-order differential equations with variable coefficients. Factorization of the sixth-order operators into second-order operators is discussed, and the details of solution are worked out for the case in which the equations are reduced to those for the isotropic shell. It is shown that equations for the latter case agree exactly with those derived previously by Reissner [Eric Reissner, “On Vibrations of Shallow Spherical Shells,” J. Appl. Phys. 17, 1038–1042 (1946)]. A table of calculated frequencies is given along with some experimental results for steel shells. (This investigation was conducted under Contract No. DA-30-115-509-ORD-912, Department of Army Ordnance Corps, Ballistic Research Laboratory, Aberdeen, Maryland.)

The paper presents a theoretical development of the equations of motion for the flexural vibrations of a shallow spherical shell of orthotropic material. For symmetrical vibrations, the motion is described by two ordinary sixth-order differential equations with variable coefficients. Factorization of the sixth-order operators into second-order operators is discussed and the details of solution are worked out for the case in which the equations are reduced to those for the isotropic shell. It is shown that equations for the latter case agree exactly with those derived previously by Reissner. A table of calculated frequencies is given along with some experimental results for steel shells.