Refined Genetic Algorithm Simulations to Model Proteins

The advent of completely sequenced genomes is leading to an unprecedented growth of sequence information while adequate structure information is often lacking. Genetic algorithm simulations have been refined and applied as a helpful tool for this question. Modified strategies are tested first on simple lattice protein models. This includes consideration of entropy (protein adjacent water shell) and improved search strategies (pioneer search +14%, systematic recombination +50% in search efficiency). Next, extension to grid free simulations of proteins in full main chain representation is examined. Our protein main chain simulations are further refined by independent criteria such as fitness per residue to judge predicted structures obtained at the end of a simulation. Protein families and protein interactions predicted from the complete H. pylori genomic sequence demonstrate how the full main chain simulations are then applied to model new protein sequences and protein families apparent from genome analysis.     

Outer surface protein of bacteriophage lambda

The bacteriophage λ capsid is composed of a main shell protein (pE) and an outer surface protein (pD). The outer surface protein was purified from sources of free protein and assembled protein. The amino acid composition, C- and N-terminals, iso-electric point, molecular weight, and state of aggregation were determined. In vitro the outer surface protein binds specifically to structures composed of λ main shell protein in the expanded configuration i.e. to enlarged preheads, pD-deficient bacteriophage particles, and polyheads.We discuss the binding of pD to the shell surface as a “pseudo-crystallisation process”, its clustering on the surface as trimers and its role as stabiliser of the filled head.     

Electrophoretic patterns of cell wall protein as a criterion for the identification and classification of Corynebacteria

Electrophoretic patterns of cell wall protein of three industrial strains, that were used for production of lysin, and eight collection strains from the genus Corynevacterium were studied to analyze their similarity as well as to estimate an opportunity of using this parameter as an additional criterion for identification and classification of corynebacteria. Similarity coefficient of cell wall overall and main protein electrophoretic patterns were determined by a specially created computer program. Electrophoretic analysis showed that every specie had an individual protein profile. There were determined biopolymers common for the specie, genus and individual among the overall majors and minors. The obtained results showed, that the patterns of main proteins were more conservative and informative in comparison with those ones of overall proteins. The definition of similarity coefficient by the main protein patterns has correlated with the protein profile characteristics of every analyzed strain, and it managed to distribute them into the separate groups. The similarity coefficient of preparations by the main protein patterns allows to separate one specie or a strain from another, and that gives us a chance to claim that this parameter could be used as an additional criterion for differentiation and referring the corynebacteria to a certain taxonomic group.     

On basic proteins in bovine peripheral nerve myelin.

1. Myeline proteins in bovine peripheral nerve migrated as two main band-(BF and BR protein) and one faint middle band (BM protein) on sodium dodecyls sulfate-polyacrylamide gel electrophoresis. The relative mobility of these two main bands differed from those of myelin proteins in the central nervous system. 2. The acid extract of the myelin fraction from bovine peripheral nerve was separated into one main peak and two minor peaks on a Sephadex G-75 column. The major component of the second minor peak was the BM protein; the major component of the main peak was the BF protein. The BR protein was not extractable by acid solution. 3. Molecular weights of the BF, the BM and the BR protein were determined as around 13 000, 20 000 and 28 000, respectively, by sodium dodecylsulfate-polyacrylamide gel electrophoresis. 4. The amino acid composition of the BF protein was quite different from the encephalitogenic protein and the Folch-Lees type proteolipid protein in the central nervous system. However the BM protein showed similar amino acid composition to the encephalitogenic protein. 5. The tryptic peptide maps of the BF protein and of the encephalitogenic protein were quite different. The results suggested that the amino acid sequences of these two proteins are different and that they contain no common tryptophan-containing peptide.     

Identification of the inhibitor of the plasminogen activator as the major protein secreted by endothelial rat liver cells

Freshly isolated Kupffer and endothelial liver cells exhibit a rate of 'de novo' protein synthesis which is twice as high per mg cell protein as that of parenchymal liver cells and contribute significantly (7.5% and 5.9%, respectively) to total liver protein secretion. In parenchymal cells the main secretory protein is a 68 kDa protein (containing 19% fo the secreted radioactivity, presumably albumin). In Kupffer cells a 49 kDa protein contains 8% of the secreted radioactivity, while in endothelial liver cells a 55 kDa protein is the most prominent secretory protein (containing 11% of the secreted radioactivity). By aid of a specific antibody the 55 kDa protein was identified as the inhibitor of the plasminogen activator and in the liver this protein was only secreted by the endothelial cells.     

Invariable chain (Ii-chain) as a highly important molecular chaperone of histocompatibility class II molecules

The publications on the structural and functional features of the main molecular shaperon of li-chain were updated. In the 1990s the determination of molecular shaperon as a group of mutually unrelated protein molecules taking part in the assembly of other polypeptides was worked out. In humans the main isoform of li-chain is the protein with a mol. wt. of 31-33 kD. A great variety of functions of this shaperon is linked with definite amino acid sequences. In particular, the molecular shaperon of the molecules of the main histocompatibility complex-II has functions connected with the presentation of antigen and the differentiation of B lymphocytes.     

Membrane separation of protein precipitates: Studies with cross flow in hollow fibers

Hollow fiber ultrafiltration and microfiltration membranes are examined for the processing of isoelectric soya protein precipitate suspensions. A model based on the various resistances to permeate flux is used to describe membrane performance. The main resistance to permeate flux is due to the interaction between the active membrane and the soluble and precipitated protein; that is, as compared with resistances due to the active membrane itself or the membrane support structure, or arising from concentrated soluble or precipitated protein layers over the membrane surface. Soluble protein rejection and precipitate mean particle diameter are correlated with observed values of this main resistance.In contract to the ultrafiltration of soluble proteins, the flux rates observed when processing protein precipitate suspensions under a similar range of operating conditions do not approach a limiting value with increased transmembrane pressure. At high protein concentrations, greater flux rates may be achieved for precipitated as compared with soluble proteins. The use of a microfiltration membrane does not give further improvement in flux rate; this may be attributed to problems of pore fouling with precipitate particles.     

The heterogeneity of protein AA in secondary (reactive)systemic amyloidosis

In secondary systemtic amyloidosis, amyloid fibrils have protein AA as a main subunit protein. As judged from gel chromatography and electrophoresis, this protein is rather homogeneous. In the present paper it is shown, however, that protein AA is very heterogeneous and composed of many peptides with different isoelectric points. However, their antigenic properties and amino acid compositions vary only little. It is concluded that protein AA is as heterogeneous as its postulated precursor, the acute phase reactant serum AA and that a theory that only one or a few serum protein AA's can give rise to amyloid fibrils, might be wrong.     

Poly(ADP-ribose) polymerase-1: A Regulator of Protein–Nucleic Acid Interactions in Processes Responding to Genotoxic Impact

Poly(ADP-ribose) polymerase-1 (PARP-1), a nuclear protein of higher eukaryotes, specifically detects strand breaks in DNA. The enzyme is activated in the presence of such breaks and synthesizes poly(ADP-ribose) covalently bound to certain proteins, with PARP-1 itself being the main acceptor. This protein is involved in the majority of DNA-dependent processes, including replication, recombination, repair, and cell death (apoptosis and necrosis). Poly(ADP-ribosyl)ation of proteins is regarded as a mechanism which induces a signal of DNA damage and modulates the function of proteins in response to genotoxic actions. Attention in this review is focused on the role of PARP-1 and poly(ADP-ribosyl)ation in base excision repair (BER), the main process of DNA break repair. The main putative functions of PARP-1 in this process are also considered, namely, its functions as a factor initiating the BER protein complex, a temporary protector of DNA ends, a factor modulating chromatin structure through poly(ADP-ribosyl)ation of histones, and a signal in the mechanism recognizing the degree of DNA damage in the cell.     

Efficient use of lactose for the lac promotercontrolled overexpression of the main antigenic protein of the foot and mouth disease virus in Escherichia coli under fed-batch fermentation conditions

Abstract: Derivatives of the lac promoter (tac, pac, rac) belong to the strongest bacterial promoters which are frequently used for the induced overexpression of foreign genes in Escherichia coli . However, their use in fermentation processes is strongly restricted because of the high cost of the inducer iso-propyl-β-D-thiogalactopyranoside (IPTG). The aim of this work was to investigate the possibility of using lac-derived promoters in high cell density processes resulting in a high yield of the induced recombinant protein if glucose is the main carbon and energy source. Lactose is tested as inducer of the main antigenic coat protein (VP1) of the foot and mouth disease (FMD) virus in a T7-RNA polymerase expression system. It was shown that lactose is able to induce the expression of the recombinant gene to an amount of the VP1 protein corresponding to 20% of the total cell protein.     

Poly(ADP-ribose) polymerase-1 as a regulator of protein-nucleic acid interactions in the processes responding to genotoxic action

Poly(ADP-ribose) polymerase-1 (PARP-1), nuclear protein of higher eukaryotes, specifically detects strand breaks in DNA. When bound to DNA strand breaks, PARP-1 is activated and catalyzes synthesis of poly(ADP-ribose) covalently attached to the row of nuclear proteins, with the main acceptor being PARP-1 itself. This protein participates in a majority of DNA dependent processes: repair, recombination; replication: cell death: apoptosis and necrosis. Poly(ADP-ribosyl)ation of proteins is considered as mechanism, which signals about DNA damage and modulate protein functioning in response to genotoxic impact. The main emphasis is made on the roles of PARP-1 and poly(ADP-ribosyl)ation in base excision repair (BER), the process, which provides repair of DNA breaks. The main proposed functions of PARP-1 in this process are: factor initiating assemblage of protein complex of BER; temporary protection of DNA ends; modulation of chromatin structure via poly(ADP-ribosyl)ation of histones; signaling function in detection of the levels of DNA damage in cell.     

Efficient unfolding pattern recognition in single molecule force spectroscopy data

Background

Single-molecule force spectroscopy (SMFS) is a technique that measures the force necessary to unfold a protein. SMFS experiments generate Force-Distance (F-D) curves. A statistical analysis of a set of F-D curves reveals different unfolding pathways. Information on protein structure, conformation, functional states, and inter- and intra-molecular interactions can be derived.

Results

In the present work, we propose a pattern recognition algorithm and apply our algorithm to datasets from SMFS experiments on the membrane protein bacterioRhodopsin (bR). We discuss the unfolding pathways found in bR, which are characterised by main peaks and side peaks. A main peak is the result of the pairwise unfolding of the transmembrane helices. In contrast, a side peak is an unfolding event in the alpha-helix or other secondary structural element. The algorithm is capable of detecting side peaks along with main peaks. Therefore, we can detect the individual unfolding pathway as the sequence of events labeled with their occurrences and co-occurrences special to bR's unfolding pathway. We find that side peaks do not co-occur with one another in curves as frequently as main peaks do, which may imply a synergistic effect occurring between helices. While main peaks co-occur as pairs in at least 50% of curves, the side peaks co-occur with one another in less than 10% of curves. Moreover, the algorithm runtime scales well as the dataset size increases.

Conclusions

Our algorithm satisfies the requirements of an automated methodology that combines high accuracy with efficiency in analyzing SMFS datasets. The algorithm tackles the force spectroscopy analysis bottleneck leading to more consistent and reproducible results.     

乳清蛋白在临床营养中的应用

乳清蛋白被认为是"蛋白之王",是人乳蛋白的主要成分。乳清蛋白的临床功用包括维持和提高机体免疫力、抗自由基延缓衰老、维持肾功能和促进创伤愈合。疾病防治作用包括防治"四高症"、心脑血管疾病和消化系统疾病,以及有助于癌症康复和AIDS患者的治疗。     

Immunological Nonidentity of 19K Protein and TP0 in Peripheral Nervous System Myelin

Peripheral nervous system myelin contains as the major structural protein a glycoprotein known as P0. Another glycoprotein present in smaller amounts, known as the 19K or X protein, has been previously identified as derived from P0 and identical with the main tryptic degradation product of P0 (TP0). Although both P0 and 19K protein incorporated fucose in vitro and stained on polyacrylamide gels with the periodic acid-Schiff stain for carbohydrate, only the P0 blotted to nitrocellulose paper showed immunoreactivity to an antibody to P0, whereas the 19K protein did not. Furthermore, when P0 was hydrolyzed with trypsin or elastase, the main degradation products reacted with P0 on immunoblots, whereas the 19K protein showed no immunoreactivity. From these studies and those of others, it may be concluded that the 19K protein shows some similarities to TP0, but probably has a different structure. P0 and 19K protein do not appear to be related as shown by lack of cross-immunoreactivity.     

Semantic similarity analysis of protein data: assessment with biological features and issues

The integration of proteomics data with biological knowledge is a recent trend in bioinformatics. A lot of biological information is available and is spread on different sources and encoded in different ontologies (e.g. Gene Ontology). Annotating existing protein data with biological information may enable the use (and the development) of algorithms that use biological ontologies as framework to mine annotated data. Recently many methodologies and algorithms that use ontologies to extract knowledge from data, as well as to analyse ontologies themselves have been proposed and applied to other fields. Conversely, the use of such annotations for the analysis of protein data is a relatively novel research area that is currently becoming more and more central in research. Existing approaches span from the definition of the similarity among genes and proteins on the basis of the annotating terms, to the definition of novel algorithms that use such similarities for mining protein data on a proteome-wide scale. This work, after the definition of main concept of such analysis, presents a systematic discussion and comparison of main approaches. Finally, remaining challenges, as well as possible future directions of research are presented.     

Determination of protein in extracts containing interfering substances and in radioactive samples following scintillation counting.

A method is described for determination of protein in biological preparations containing various interfering substances normally present in extraction media. The main steps of the procedure consist in depositing the protein solution on filter paper strips and removing all small molecular weight substances by washing with a number of aqueous and nonaqueous solvents. The protein remaining on the paper is then determined by a modification of Lowry's colorimetric procedure. The method also permits the determination of protein in radioactive samples which have been previously counted using a liquid scintillation mixture such as Apuasol or toluene-POPOP.     

Cell-free synthesis of proteins related to sn-glycerol-3-phosphate transport in Escherichia coli.

An Escherichia coli periplasmic protein (GlpT) related to sn-glycerol-3-phosphate transport was synthesized in a cell-free system directed by hybrid plasmic ColE1-glpT DNA. The in vitro product cross-reacted with antisera against the purified protein. The ColE1-glpT DNA-directed cell-free system was induced by sn-glycerol-3-phosphate and phosphonomycin and was dependent on cyclic AMP. The in vitro-synthesized protein showed the characteristics of a multimeric protein, as did the purified periplasmic protein. The main proportion of the newly synthesized product had a higher molecular weight than the mature protein found in the periplasm of cells and showed a more positive charge in two-dimensional gel electrophoresis. Thus, a proportion of this protein is presumed to be synthesized in vitro as a precursor. The cell-free system yielded a second protein that is likely to be also coded for by the glpT operon. This protein had a molecular weight of approximately 33,000 in sodium dodecyl sulfate-acrylamide gel electrophoresis and behaved like an intrinsic membrane protein.     

Emerging roles for protein S-palmitoylation in Toxoplasma biology

Post-translational modifications are refined, rapidly responsive and powerful ways to modulate protein function. Among post-translational modifications, acylation is now emerging as a widespread modification exploited by eukaryotes, bacteria and viruses to control biological processes. Protein palmitoylation involves the attachment of palmitic acid, also known as hexadecanoic acid, to cysteine residues of integral and peripheral membrane proteins and increases their affinity for membranes. Importantly, similar to phosphorylation, palmitoylation is reversible and is becoming recognised as instrumental for the regulation of protein function by modulating protein interactions, stability, folding, trafficking and signalling. Palmitoylation appears to play a central role in the biology of the Apicomplexa, regulating critical processes such as host cell invasion which is vital for parasite survival and dissemination. The recent identification of over 400 palmitoylated proteins in Plasmodium falciparum erythrocytic stages illustrates the broad spread and impact of this modification on parasite biology. The main enzymes responsible for protein palmitoylation are multi-membrane protein S-acyl transferases harbouring a catalytic Asp-His-His-Cys (DHHC) motif. A global functional analysis of the repertoire of protein S-acyl transferases in Toxoplasma gondii and Plasmodium berghei has recently been performed. The essential nature of some of these enzymes illustrates the key roles played by this post-translational modification in the corresponding substrates implicated in fundamental processes such as parasite motility and organelle biogenesis. Toward a better understanding of the depalmitoylation event, a protein with palmitoyl protein thioesterase activity has been identified in T. gondii. TgPPT1/TgASH1 is the main target of specific acyl protein thioesterase inhibitors but is dispensable for parasite survival, suggesting the implication of other genes in depalmitoylation. Palmitoylation/depalmitoylation cycles are now emerging as potential novel regulatory networks and T. gondii represents a superb model organism in which to explore their significance.