Analyzing the normal mode dynamics of macromolecules by the component synthesis method: Residue clustering and multiple-component approach

A study of the component synthesis method (CSM) for analyzing the normal mode dynamics of macromolecules is reported. The procedure involves a reduction of the dimensions of the normal mode problems for large molecular systems and the accurate extraction of the low-frequency modes. A macromolecule is divided into small components based on a hierarchical clustering of the residues in the structure. Interactions between coupled components are treated by the method of static correlation. The normal modes of the components are obtained first, and a fraction of the low-frequency normal modes of the components under mutual correlations are then used as a reduced basis for solving for the normal modes of the whole molecule. Multiple components are introduced for large macromolecules so that the dimensions of the eigenvalue problems at the component level are small. The method is applied to the protein crambin. In test calculations in which the dimensions of the eigenvalue equations are reduced to 1/6 of their natural size, the errors in the normal mode frequencies calculated by the CSM procedure are only about 1–2% when compared with the exact values. The rms fluctuations of all atoms in crambin calculated by the CSM procedure are basically identical to the exact results. The CSM procedure is shown to be accurate for calculating the normal modes of large macromolecules with a significant reduction of the size of the problem. © 1994 John Wiley & Sons, Inc.     

Gastroduodenal ulceration following active immunization with prostaglandin E2 in dogs. Role of gastric acid secretion

In this study we present evidence to suggest that gastroduodenal mucosal defects may occur in gastric fistula dogs actively immunized with PGE2-thyroglobulin conjugate. One of four PGE2-immunized dogs developed a chronic pyloroduodenal ulcer with penetration into the pancreas and the other three had endoscopic evidence of gastric and/or duodenal erosions. In contrast, no gastroduodenal mucosal defects were seen in control dogs immunized with thyroglobulin alone. Occurrence of gastroduodenal ulcers or erosions was temporally related to formation of specific antibody to PGE2 suggesting that PGE2 antibody may be responsible for lesion formation. An increase in gastric acid secretion was not observed in PGE2-immunized dogs. Thus, it is likely that mucosal defects occur as a result of an impairment of PGE2-mediated mucosal defense mechanisms. Since gastroduodenal lesions can be visualized by endoscopy, the dog may prove to be useful in studying the role of endogenous PG in ulcer diseases.     

Private Financing Options for Long-term Care

Private financing for long-term care now comes almost exclusively from out-of-pocket payments. Long-term-care costs quickly impoverish most elderly, resulting in Medicaid dependency. The consequences are profound for the western Sun Belt with its rapidly growing elderly population. Key private financing options are long-term-care individual retirement accounts (LTC/IRAs), home equity conversion, social-health maintenance organizations and long-term-care insurance. Study of data from the past half century suggests that the LTC/IRA approach would prove unsatisfactory for the purpose despite the intuitive appeal of this mechanism. Experience with home equity conversions is still very limited, and unresolved questions limit this approach to the role of a reserve option for now. While promising, social-health maintenance organizations are still in the experimental stages and not yet commercially available. Long-term-care insurance is currently sold on a thin market and emphasizes nursing home coverage. New approaches to private financing through long-term-care insurance seem to offer the best approach for immediate implementation.     

Carbohydrate recognition systems in amphibians: primitive alpha 2-macroglobulin receptors

Two alpha-macroglobulins were isolated from the plasma of the frog. Murine clearance studies were performed with these proteins after they were reacted with proteinase. These studies indicated that clearance behavior was more complex than observed with avian or human homologues, since it involved not only specific receptors for the complex, but also carbohydrate recognition. Recognition of one of these macroglobulins by both the alpha-macroglobulin receptor and carbohydrate receptors required conformational change in the inhibitor. Clearance studies were then performed in frogs to probe the nature of carbohydrate receptor recognition. Model glycoproteins were employed to avoid the problem of heterogeneous carbohydrate end groups in the alpha-macroglobulins. These studies demonstrated that the N-acetylglucosamine/mannose receptor is the major carbohydrate recognition system in frogs.     

Product of SEC53 is required for folding and glycosylation of secretory proteins in the lumen of the yeast endoplasmic reticulum

Yeast secretory mutant sec53 cells accumulate inactive secretory glycoprotein precursors that remain associated with the endoplasmic reticulum (ER) at the restrictive temperature (37 degrees C). The possibility that precursor polypeptides fail to penetrate completely into the ER lumen was tested by examining the protease accessibility of accumulated invertase, mating pheromone precursor prepro-alpha-factor and the vacuolar protein precursor procarboxypeptidase Y in cell lysates. In all three cases, the secretory protein precursors are protected from the action of exogenous protease unless the membrane is permeabilized by including Triton X-100 or saponin in the incubation. These results suggest that the sec53 defect allows complete polypeptide translocation. Consistent with this interpretation, the precursor of invertase accumulates in a signal peptide-processed form. In addition, invertase and prepro-alpha-factor precursors contain a small amount of possibly aberrant carbohydrate. In mutant cells or in wild type cells treated with tunicamycin, a 10-kDa fragment of the N terminus of mature invertase assumes a conformation that is resistant to trypsin with or without detergent. This domain may be associated with an ER protein or may simply assume an unusual conformation as a consequence of deficient glycosyl modification.     

Purification and Properties of Rubrophilin: A Novel Brain Specific Membrane Polypeptide

Rubrophilin, a unique brain specific polypeptide, was purified to apparent homogeneity from microsomal fractions of bovine brains. The peptide stains pink with Coomassie Brilliant Blue R-250 (C.I. No. 42660) under specific conditions, has an apparent Mr of 53,000, and is acidic with an apparent pI of 4.9. The purification involves initial solubilization of delipidated microsomes in sodium dodecyl sulfate, followed by ammonium sulfate fractionation, reversed ammonium sulfate gradient elution from diatomaceous earth, gel filtration on polyacrylamide (Biogel P-200), gradient elution chromatography from hydroxylapatite, and reverse-phase chromatography from phenyl-Sepharose. A yield of about 5 mg of rubrophilin was obtained from 9 g of microsomal proteins. Amino acid analysis shows that rubrophilin contains only nine amino acids with residues/mol as follows: alanine (102), glutamic acid (97), lysine (65), proline (55), aspartic acid (48), glycine (44), serine (37), threonine (35), and valine (10). Cysteine, methionine, tryptophan, tyrosine, isoleucine, phenylalanine, histidine, and arginine could not be detected. Relative rubrophilin content of vertebrate brains was as follows: mammals greater than birds greater than reptiles greater than fishes. It is present in mouse retina and human neuroblastoma cell cultures but could not be detected in octopus optic lobe or in cultured C-6 rat glioma cells.