Five Hierarchical Levels of Sequence-Structure Correlation in Proteins

This article reviews recent work towards modelling protein folding pathways using a bioinformatics approach. Statistical models have been developed for sequence-structure correlations in proteins at five levels of structural complexity: (i) short motifs; (ii) extended motifs; (iii) nonlocal pairs of motifs; (iv) 3-dimensional arrangements of multiple motifs; and (v) global structural homology. We review statistical models, including sequence profiles, hidden Markov models (HMMs) and interaction potentials, for the first four levels of structural detail. The I-sites (folding Initiation sites) Library models short local structure motifs. Each succeeding level has a statistical model, as follows: HMMSTR (HMM for STRucture) is an HMM for extended motifs; HMMSTR-CM (Contact Maps) is a model for pairwise interactions between motifs; and SCALI-HMM (HMMs for Structural Core ALIgnments) is a set of HMMs for the spatial arrangements of motifs. The parallels between the statistical models and theoretical models for folding pathways are discussed in this article; however, global sequence models are not discussed because they have been extensively reviewed elsewhere. The data used and algorithms presented in this article are available at http://www.bioinfo.rpi.edu/~bystrc/ (click on "servers" or "downloads") or by request to bystrc@rpi.edu .     

Male mouse meiotic chromosome cores deficient in structural proteins SYCP3 and SYCP2 align by homology but fail to synapse and have possible impaired specificity of chromatin loop attachment

The targeted deletion of the meiotic chromosome core component MmSYCP3 results in chromosome synaptic failure at male meiotic prophase, extended meiotic chromosomes, male sterility, oocyte aneuploidy and absence of the MmSYCP2 chromosome core component. To test the functions of SYCP2 and SYCP3 proteins in the cores, we determined the effect of their deletion on homology recognition by whole chromosome painting and the effect on chromatin loop attachment to the cores with endogenous and exogenous sequences. Because we observed that the alignment of cores is between homologs, it suggested that alignment is not a function of the chromosome core components but might be mediated by chromatin-chromatin interactions. The alignment function therefore appears to be separate from intimate synapsis function of homologous cores that is observed to be defective in the SYCP3-/- males. To examine the functions of the SYCP2 and 3 core proteins in chromatin loop attachment, we measured the loop sizes of the centromeric major satellite chromatin and the organization of an exogenous transgene in SYCP3+/+ and SYCP3-/- males. We observed that these satellite chromatin loops have a normal appearance in SYCP3-/- males, but the loop regulation of a 2-Mb exogenous lambda phage insert appears to be altered. Normally the insert fails to attach to the core except by flanking endogenous sequences, but in the absence of SYCP2 and SYCP3, there appears to be multiple attachments to the core. This suggests that the selective preference for the attachment of mouse sequences to the chromosome core in the wild-type male is impaired in the SYCP3-/- male. Apparently the SYCP2 and SYCP3 proteins function in the specificity of chromatin attachment to the chromosome core.     

Intein-mediated rapid purification of recombinant human pituitary adenylate cyclase activating polypeptide

In order to obtain the recombinant human PACAP efficiently by intein-mediated single column purification, a gene encoding human PACAP was synthesized and cloned into Escherichia coli expression vector pKYB. The recombinant vector pKY-PAC was transferred into E. coli ER2566 cells and the target protein was over-expressed as     

Expression of a modified Cry1Ie gene in E. coli and in transgenic tobacco confers resistance to corn borer

The wild-type Crylle gene from Bacillus thuringiensis was modified for its efficient expression in transgenic plants. Modified Crylle gene (designated as Cryllem) was cloned into prokaryotic expressionvector pET28b and its expression in E.coli was confirmed by SDS-PAGE analysis. Bioassays using crude expression products in E.coli revealed that CrylIem protein had a similar toxicity to corn borer as wild-type CrylIe. CrylIem gene was then inserted downstream of the maize ubiquitin-1 promoter in plant expression vector p3301. Transgenic tobacco plants carrying Cryllem showed insecticidal activity against corn borer.     

SCOPmap: Automated assignment of protein structures to evolutionary superfamilies

Background  

Inference of remote homology between proteins is very challenging and remains a prerogative of an expert. Thus a significant drawback to the use of evolutionary-based protein structure classifications is the difficulty in assigning new proteins to unique positions in the classification scheme with automatic methods. To address this issue, we have developed an algorithm to map protein domains to an existing structural classification scheme and have applied it to the SCOP database.     

Inclusion complexes of paclitaxel and oligo(ethylenediamino) bridged bis(beta-cyclodextrin)s: solubilization and antitumor activity

The inclusion complexation behavior of paclitaxel with a series of oligo(ethylenediamino) bridged bis(beta-cyclodextrin)s possessing bridge chains in different length (1-4) has been investigated in order to improve the water solubility of paclitaxel. It is found that only the long-tethered bis(beta-cyclodextrin)s 1 and 2 can form the inclusion complexes with paclitaxel, which are characterized by NMR, SEM, XRD, FT-IR, TG-DTA, DSC, and microcalorimetry technology. The results obtained show that bis(beta-cyclodextrin)s 1 and 2 are able to solubilize paclitaxel to high levels up to 2 and 0.9 mg/mL, respectively. The high complex stability of bis(beta-cyclodextrin) 1 and paclitaxel is discussed from thermodynamic viewpoint. Furthermore, the cytotoxicity of these complexes assessed using a human erythroleukemia K562 cell line indicates that the IC(50) value of 1/paclitaxel complex is 6.0 x 10(-10) mol/dm(3) (calculated as paclitaxel molar concentration), which means that the antitumor activity of 1/paclitaxel complex is better than that of parent paclitaxel (IC(50) value 9.8 x 10(-10) mol/dm(3)). This high antitumor activity, along with the satisfactory water solubility and high thermal stability of the 1/paclitaxel complex, will be potentially useful for its clinical application as a highly effective antitumor drug.     

The cell cycle related apoptotic susceptibility to arsenic trioxide is associated with the level of reactive oxygen species

Double staining flow cytometry was performed using 7-amino actinomycin D and 6-carboxy-2‘,7‘-dichlorodihydrofluorescein diacetate, to detect the level fluctuation of reactive oxygen species (ROS) during the cell cycle of normal NB4 cells. Our results showed that NB4 cells possessed higher level of ROS in G2/M phase than in G1 and S phases. Double staining flow cytometry, with TdT mediated dUTP nick end labeling (Tunel) and propidium iodide(PI), indicated that As2O3 (2μM) could induce apoptosis in NB4 cells prevailingly from G2/M phase, and this efficacy was enhanced upon co-administration of 2, 3-dimethoxy-1, 4-naphthoquinone (DMNQ) (2.5μM) which could produce the endogenous ROS. These results suggested that different ROS level in different cell cycle phases of NB4 cells might determin the selective induction of G2/M apoptosis and the cells‘ susceptibility to apoptosis by As2O3.