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91.
92.
Diekman C He W Prabhu N Cramer H 《Analytical and quantitative cytology and histology / the International Academy of Cytology [and] American Society of Cytology》2003,25(4):183-190
OBJECTIVE: To design and analyze a new family of hybrid methods for the diagnosis of breast tumors using fine needle aspirates. STUDY DESIGN: We present a radically new approach to the design of diagnosis systems. In the new approach, a nonlinear classifier with high sensitivity but low specificity is hybridized with a linear classifier having low sensitivity but high specificity. Data from the Wisconsin Breast Cancer Database are used to evaluate, computationally, the performance of the hybrid classifiers. RESULTS: The diagnosis scheme obtained by hybridizing the nonlinear classifier ellipsoidal multisurface method (EMSM) with the linear classifier proximal support vector machine (PSVM) was found to have a mean sensitivity of 97.36% and a mean specificity of 95.14% and was found to yield a 2.44% improvement in the reliability of positive diagnosis over that of EMSM at the expense of 0.4% degradation in the reliability of negative diagnosis, again compared to EMSM. At the 95% confidence level we can trust the hybrid method to be 96.19-98.53% correct in its malignant diagnosis of new tumors and 93.57-96.71% correct in its benign diagnosis. CONCLUSION: Hybrid diagnosis schemes represent a significant paradigm shift and provide a promising new technique to improve the specificity of nonlinear classifiers without seriously affecting the high sensitivity of nonlinear classifiers. 相似文献
93.
Cramer H Rhea R Okicki J Yirava K Silverman R 《Nucleosides, nucleotides & nucleic acids》2003,22(5-8):1733-1736
The 2-5A antisense compound RBI 011 targeting telomerase RNA was synthesized using the triisopropylsilyl-oxymethyl (TOM) group for the 3'-hydroxyl protection of 2',5'-linked RNA. 相似文献
94.
Ponamarev MV Schlarb BG Howe CJ Carrell CJ Smith JL Bendall DS Cramer WA 《Biochemistry》2000,39(20):5971-5976
Cytochrome f of oxygenic photosynthesis has an unprecedented structure, including the N-terminus being a heme ligand. The adjacent N-terminal heme-shielding domain is enriched in aromatic amino acids. The atomic structures of the chloroplast and cyanobacterial cytochromes f were compared to explain spectral and redox differences between them. The conserved aromatic side chain in the N-terminal heme-shielding peptide at position 4, Phe and Tyr in plants and algae, respectively, and Trp in cyanobacteria, is in contact with the heme. Mutagenesis of cytochrome f from the eukaryotic green alga Chlamydomonas reinhardtii showed that a Phe4 --> Trp substitution in the N-terminal domain was unique in causing a red shift of 1 and 2 nm in the cytochrome Soret (gamma) and Q (alpha) visible absorption bands, respectively. The resulting alpha band peak at 556 nm is characteristic of the cyanobacterial cytochrome. Conversely, a Trp4 --> Phe mutation in the expressed cytochrome from the cyanobacterium Phormidium laminosum caused a blue shift to the 554 nm alpha band peak diagnostic of the chloroplast cytochrome. Residue 4 was found to be the sole determinant of this 60 cm(-)(1) spectral shift, and of approximately one-half of the 70 mV redox potential difference between cytochrome f of P. laminosum and C. reinhardtii (E(m7) = 297 and 370 mV, respectively). The proximity of Trp-4 to the heme implies that the spectral and redox potential shifts arise through differential interaction of its sigma- or pi-electrostatic potential with the heme ring and of the pi-potential with the heme Fe orbitals, respectively. The dependence of the visible spectrum and redox potential of cytochrome f on the identity of aromatic residue 4 provides an example of the use of the relatively sharp cytochrome spectrum as a "spectral fingerprint", and of the novel structural connection between the heme and a single nonliganding residue. 相似文献
95.
The colicin E1 immunity protein (ImmE1), a 13.2-kDa hydrophobic integral membrane protein localized in the Escherichia coli cytoplasmic membrane, protects the cell from the lethal, channel-forming activity of the bacteriocin, colicin E1. Utilizing its solubility in organic solvents, ImmE1 was purified by 1-butanol extraction of isolated membranes, followed by gel filtration and ion-exchange chromatography in a chloroform/methanol/H(2)O (4:4:1) solvent system. Circular dichroism analysis indicated that the alpha-helical content of ImmE1 is approximately 80% in 1-butanol or 2,2,2-trifluoroethanol, consistent with a previous membrane-folding model with three extended hydrophobic transmembrane helical domains, H1-H3. Each of these extended hydrophobic domains contains a centrally located single Cys residue that could be used as a probe of protein structure. The presence of tertiary structure of purified ImmE1 in a solvent of mixed polarity, chloroform/methanol/H(2)O (4:4:1) was demonstrated by (i) the constraints on Tyr residues shown by the amplitude of near-UV circular dichroism spectra in the wavelength interval, 270-285 nm; (ii) the correlation between the near-UV Tyr CD spectrum of single and double Cys-to-X mutants of the Imm protein and their in vivo activity; (iii) the upfield shift of methyl groups in a 1D NMR spectrum, a 2D- HSQC NMR spectrum of ImmE1 in the mixed polarity solvent mixture, and a broadening and disappearance of the indole (1)H proton resonance from Trp94 in H3 by a spin label attached to Cys16 in the H2 hydrophobic domain; (iv) near-UV circular dichroism spectra with a prominent ellipticity band centered at 290 nm from a single Trp inserted into the extended hydrophobic domains. It was concluded that the colicin E1 immunity protein adopts a folded conformation in chloroform/methanol/H(2)O (4:4:1) that is stabilized by helix-helix interactions. Analysis of the probable membrane folding topology indicated that several Tyr residues in the bilayer region of the three transmembrane helices could contribute to the near-UV CD spectrum through helix-helix interactions. 相似文献
96.
Displacement chromatography of proteins was successfully carried out in both hydrophobic interaction and reversed-phase chromatographic systems using low-molecular weight displacers. The displacers employed for hydrophobic displacement chromatography were water soluble, charged molecules containing several short alkyl and/or aryl groups. Spectroscopy was employed to verify the absence of structural changes to the proteins displaced on these hydrophobic supports. Displacement chromatography on a reversed-phase material was employed to purify a growth factor protein from its closely related variants, demonstrating the high resolutions that can be achieved by hydrophobic displacement chromatography. This process combines the high-resolution/high-throughput characteristics of displacement chromatography with the unique selectivity of these hydrophobic supports and offers the chromatographic engineer a powerful tool for the preparative purification of proteins. 相似文献
97.
Dabkowski W Tworowska I Michalski J Cramer F 《Nucleosides, nucleotides & nucleic acids》2000,19(10-12):1779-1785
We describe the convenient synthesis of thymidine cyclic 3', 5'-phosphorofluoridate 6, which is superior to that previously reported. Our procedure is based on a sequence of reactions utilizing 3 as the key substrate. Similar sequence of reaction leads to the sulfur analogues of 6 the thymidine cyclic 3',5'-phosphorofluoridothioate 7. 相似文献
98.
99.
By replacing two cysteine residues in apamin with selenocysteine, the three possible isomers related to the side-chain connectivities of a bis-cystinyl-peptide were synthesized in regioselective manner exploiting the low redox potential of the diselenide bond. Nuclear magnetic resonance conformational analysis of monoselenocystine analogue apamin with the natural diselenide/disulfide network confirmed the highly isomorphous character of the sulfur replacement with selenium despite its slightly larger atomic radius and increased bond lengths. The comparative conformational analysis of the apamin analogues containing the non-natural side-chain links with wild type apamin clearly revealed retention of the main structural fold and thus the high propensity of these small molecules to adopt the secondary structure elements present in natural apamin. These findings offered interesting hints for a better understanding of the oxidative refolding pathway of the bis-cystinyl peptide that leads exclusively to the correct natural isomer. 相似文献
100.
Coupled plasmon-waveguide resonance spectroscopy studies of the cytochrome b6f/plastocyanin system in supported lipid bilayer membranes. 下载免费PDF全文
The incorporation of cytochrome (cyt) b6f into a solid-supported planar egg phosphatidylcholine (PC) bilayer membrane and complex formation with plastocyanin have been studied by a variant of surface plasmon resonance called coupled plasmon-waveguide resonance (CPWR) spectroscopy, developed in our laboratory. CPWR combines greatly enhanced sensitivity and spectral resolution with direct measurement of anisotropies in refractive index and optical extinction coefficient, and can therefore probe structural properties of lipid-protein and protein-protein interactions. Cyt b6f incorporation into the membrane proceeds in two stages. The first occurs at low protein concentration and is characterized by an increase in total proteolipid mass without significant changes in the molecular order of the system, as demonstrated by shifts of the resonance position to larger incident angles without changing the refractive index anisotropy. The second stage, occurring at higher protein concentrations, results in a decrease in both the mass density and the molecular order of the system, evidenced by shifts of the resonance position to smaller incident angles and a large decrease in the membrane refractive index anisotropy. Plastocyanin can bind to such a proteolipid system in three different ways. First, the addition of plastocyanin before the second stage of b6f incorporation begins results in complex formation between the two proteins with a KD of approximately 10 microM and induces structural changes in the membrane that are similar to those occurring during the second stage of complex incorporation. The addition of larger amounts of plastocyanin under these conditions leads to nonspecific binding to the lipid phase with a KD of approximately 180 microM. Finally, the addition of plastocyanin after the completion of the second phase of b6f incorporation results in tighter binding between the two proteins (KD approximately 1 microM). Quantitation of the binding stoichiometry indicates that two plastocyanin molecules bind tightly to the dimeric form of the cyt b6f complex, assuming random insertion of the cytochrome into the bilayer. The structural basis for these results and formation of the proteolipid membrane are discussed. 相似文献