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141.
The lens influences retinal growth and differentiation during vertebrate eye development but the mechanisms are not understood. The role of the lens in retinal growth and development was studied in the teleost Astyanax mexicanus, which has eyed surface-dwelling (surface fish) and blind cave-dwelling (cavefish) forms. A lens and laminated retina initially develop in cavefish embryos, but the lens dies by apoptosis. The cavefish retina is subsequently disorganized, apoptotic cells appear, the photoreceptor layer degenerates, and retinal growth is arrested. We show here by PCNA, BrdU, and TUNEL labeling that cell proliferation continues in the adult cavefish retina but the newly born cells are removed by apoptosis. Surface fish to cavefish lens transplantation, which restores retinal growth and rod cell differentiation, abolished apoptosis in the retina but not in the RPE. Surface fish lens deletion did not cause apoptosis in the surface fish retina or affect RPE differentiation. Neither lens transplantation in cavefish nor lens deletion in surface fish affected retinal cell proliferation. We conclude that the lens acts in concert with another optic component, possibly the RPE, to promote retinal cell survival. Accordingly, deficiency in both optic structures may lead to eye degeneration in cavefish. 相似文献
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143.
Synopsis High-speed cinematography and video using modified Schlieren optics and laser illumination helped elicit details of prey capture mechanisms used by Chromis viridis while feeding on calanoid copepods and Artemia. Chromis viridis is capable of a ram-jaw, low-suction feeding, as well as a typical suction feeding behavior described for other species of planktivores. By adjusting the degree of jaw protrusion and amount of suction used during a feeding strike, this fish can modulate its feeding strikes according to the prey type being encountered. The ram-jaw feeding mode enables C. viridis to capture highly evasive calanoid copepods within 6 to 10 msec. The use of specialized feeding behavior for evasive prey and the ability to vary feeding behavior are adaptations for feeding on evasive prey. 相似文献
144.
Utility of the gas-phase sequencer for both liquid- and solid-phase degradation of proteins and peptides at low picomole levels 总被引:6,自引:0,他引:6
James E. Strickler Michael W. Hunkapiller Kenneth J. Wilson 《Analytical biochemistry》1984,140(2):553-566
The utility of the commercially available gas-phase sequencer for complete analysis of peptide samples was investigated. Using the program supplied with the instrument, significant extractive loss of samples in Polybrene was observed, even at input levels up to 500 pmol. In order to reduce this loss, the sequencer program was modified by increasing the phenylisothiocyanate (PITC)-coupling steps from two to three and lengthening the duration of ethyl acetate (S2) delivery while reducing the delivery rate. These changes gave improved results with peptides, e.g., all eight residues of angiotensin II were identified at the 25-pmol level. In addition, background contamination was decreased and repetitive yields were increased. The instrument was also found to function well with samples coupled to solid supports; however, some of the methodologies that work adequately for covalent attachment of peptides to solid supports at the level 1-10 nmol were found to give unacceptable coupling/sequenceable yields at or below the 100-pmol level. The coupling methods tried were (1) reaction of homoserine lactone with aminopropyl (AP)-glass, (2) reaction of alpha- and epsilon-NH2 groups with p-phenylenediisothiocyanate (DITC)-glass, and (3) reaction of alpha-COOH groups with aminoaryl (AA)-glass via EDAC (1-ethyl-3,3'-dimethylaminopropyl-carbodiimide). Of these, the first method gave combined yields of 42-94% while the latter two were only 9-35% efficient. The covalently bound samples provided sequence information even at the resulting low levels, e.g., 9/13 residues of dynorphin including Lys-13 at 11 pmol. In general, sequencer runs on solid-phase samples gave "cleaner" analyses and slightly higher repetitive yields (1-2%). Sequence information has also been obtained on peptides made by solid-phase synthesis prior to cleavage from the polystyrene support. With improved coupling efficiencies, solid-phase techniques would provide an alternative to immobilization of peptides in Polybrene films for low picomole level gas-phase sequencing. 相似文献
145.
Specific proteolytic modification of creatine kinase isoenzymes. Implication of C-terminal involvement in enzymic activity but not in subunit-subunit recognition. 总被引:4,自引:1,他引:3 下载免费PDF全文
H G Lebherz T Burke J E Shackelford J E Strickler K J Wilson 《The Biochemical journal》1986,233(1):51-56
We are using the isoenzymes of creatine kinase (CK) to investigate the effect of specific proteolytic modification on the abilities of enzyme subunits to establish precise subunit-subunit recognition in vitro. Previous work by others has shown that treatment of the MM isoenzyme of rabbit CK with Proteinase K results in a specific proteolytic modification and inactivation of the enzyme. In the present work, we show that both the MM and BB isoenzymes of chicken CK are also specifically modified by Proteinase K, resulting in over 98% loss of catalytic activity and approx. 10% decreases in subunit molecular masses of the enzymes. Similar reactions appear to occur when the isoenzymes are treated with Pronase E. Limited amino acid sequence analysis of intact and Proteinase K-modified MM-CK suggests that the proteolytic modification results from a single peptide-bond cleavage occurring between alanine residues 328 and 329, about 50 amino acid residues from the C-terminal end; the active-site cysteine residue was recovered in the large protein fragment of modified M-CK subunits. Proteolytically modified M-CK and B-CK subunits were able to refold and reassociate into dimeric structures after treatment with high concentrations of LiCl and at low pH. Thus the proteolytically modified CK subunits retain their ability to refold and to establish precise subunit-subunit recognition in vitro. 相似文献
146.
Characterization of the promoter, signal sequence, and amino terminus of a secreted beta-galactosidase from "Streptomyces lividans" 总被引:13,自引:11,他引:2 下载免费PDF全文
T Eckhardt J Strickler L Gorniak W V Burnett L R Fare 《Journal of bacteriology》1987,169(9):4249-4256
The gene for a secreted 130-kilodalton beta-galactosidase from "Streptomyces lividans" has been cloned, its promoter, signal sequence, and amino terminal region have been localized, and their nucleotide sequence has been determined. The signal sequence extends over 56 amino acids and shows the characteristic-features of signal sequences, including a hydrophilic amino terminus followed by a hydrophobic core near the signal cleavage site. The secretion of beta-galactosidase depends on the presence of the signal sequence. beta-Galactosidase is the major protein in culture supernatants and extracts of strains expressing the cloned beta-galactosidase gene and represents a valuable tool in the study of protein secretion in Streptomyces spp. 相似文献
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148.
A plasma membrane-enriched vesicle fraction has been prepared from Trypanosoma brucei by sonication and differential centrifugation on sucrose gradients. This fraction is enriched 5-fold in the plasma membrane marker enzymes adenyl cyclase (EC 4.6.1.1) and ouabain-inhibitable, ()-dependent adenosine triphosphatase (EC 3.6.1.3). It is also enriched up to 14-fold in iodinated surface proteins, and up to 4-fold in [3H]mannose-labeled glycoproteins, of which the major variable surface coat glycoprotein is the main constituent. Proteins of the plasma membrane fraction and other subcellular fractions have been identified by electrophoretic analysis in sodium dodecyl sulfate-polyacrylamide gradient slab gels. Several high molecular weight surface glycopeptides have been selectively investigated and partially characterized by a combination of metabolic labeling with [3H]mannose, lactoperoxidase-catalyzed surface iodination, and affinity chromatography on Con A-Sepharose. In addition to the major variable surface coat glycoprotein (estimated ), there are several minor surface glycopeptides () which are apparent extrinsic membrane components, and two surface glycopeptides () which are intrinsic membrane components. 相似文献
149.
Peptide purification via high-performance liquid chromatography (HPLC) and solid-phase sequencing were integrated to form a system allowing the determination of complete sequence information on a microscale without the use of radiolabels or modified phenylisothiocyanate. Mixtures of peptides (500 pmol to 10 nmol) resulting from proteolytic digestion or chemical cleavage were applied directly to reverse-phase columns. The columns, equilibrated in either 10 mm KPi or 0.05% trifluoroacetic acid, were then developed using acetonitrile gradients. Eluates were monitored nondestructively by direct ultraviolet detection at both 214 and 254 nm. Each peak was collected as a discrete fraction, and purity was assessed by amino acid analysis prior to covalent attachment to a solid support for sequence analysis. Activation of the peptide carboxyl terminus via a water soluble carbonyldiimide was the solid-phase coupling method used 90% of the time. Coupling yields averaged 52% of starting material. Sequence analysis was performed in the range 100 pmol to 4 nmol of coupled peptide. Phenylthiohydantoin-amino acids were identified by reverse-phase HPLC using ultraviolet detection. 相似文献
150.