GPCR-OKB: the G protein coupled receptor oligomer knowledge base

Rapid expansion of available data about G Protein Coupled Receptor (GPCR) dimers/oligomers over the past few years requires an effective system to organize this information electronically. Based on an ontology derived from a community dialog involving colleagues using experimental and computational methodologies, we developed the GPCR-Oligomerization Knowledge Base (GPCR-OKB). GPCR-OKB is a system that supports browsing and searching for GPCR oligomer data. Such data were manually derived from the literature. While focused on GPCR oligomers, GPCR-OKB is seamlessly connected to GPCRDB, facilitating the correlation of information about GPCR protomers and oligomers

Functional Diversity and Structural Disorder in the Human Ubiquitination Pathway

The ubiquitin-proteasome system plays a central role in cellular regulation and protein quality control (PQC). The system is built as a pyramid of increasing complexity, with two E1 (ubiquitin activating), few dozen E2 (ubiquitin conjugating) and several hundred E3 (ubiquitin ligase) enzymes. By collecting and analyzing E3 sequences from the KEGG BRITE database and literature, we assembled a coherent dataset of 563 human E3s and analyzed their various physical features. We found an increase in structural disorder of the system with multiple disorder predictors (IUPred - E1: 5.97%, E2: 17.74%, E3: 20.03%). E3s that can bind E2 and substrate simultaneously (single subunit E3, ssE3) have significantly higher disorder (22.98%) than E3s in which E2 binding (multi RING-finger, mRF, 0.62%), scaffolding (6.01%) and substrate binding (adaptor/substrate recognition subunits, 17.33%) functions are separated. In ssE3s, the disorder was localized in the substrate/adaptor binding domains, whereas the E2-binding RING/HECT-domains were structured. To demonstrate the involvement of disorder in E3 function, we applied normal modes and molecular dynamics analyses to show how a disordered and highly flexible linker in human CBL (an E3 that acts as a regulator of several tyrosine kinase-mediated signalling pathways) facilitates long-range conformational changes bringing substrate and E2-binding domains towards each other and thus assisting in ubiquitin transfer. E3s with multiple interaction partners (as evidenced by data in STRING) also possess elevated levels of disorder (hubs, 22.90% vs. non-hubs, 18.36%). Furthermore, a search in PDB uncovered 21 distinct human E3 interactions, in 7 of which the disordered region of E3s undergoes induced folding (or mutual induced folding) in the presence of the partner. In conclusion, our data highlights the primary role of structural disorder in the functions of E3 ligases that manifests itself in the substrate/adaptor binding functions as well as the mechanism of ubiquitin transfer by long-range conformational transitions. © 2013 Bhowmick et al

On The Impact of Internet Naming Evolution: Deployment, Performance, and Security Implications

As one of the most critical components of the Internet, the Domain Name System (DNS) provides naming services for Internet users, who rely on DNS to perform the translation between the domain names and network entities before establishing an In- ternet connection. In this dissertation, we present our studies on different aspects of the naming infrastructure in today’s Internet, including DNS itself and the network services based on the naming infrastructure such as Content Delivery Networks (CDNs). We first characterize the evolution and features of the DNS resolution in web ser- vices under the emergence of third-party hosting services and cloud platforms. at the bottom level of the DNS hierarchy, the authoritative DNS servers (ADNSes) maintain the actual mapping records and answer the DNS queries. The increasing use of upstream ADNS services (i.e., third-party ADNS-hosting services) and Infrastructure-as-a-Service (IaaS) clouds facilitates the deployment of web services, and has been fostering the evo- lution of the deployment of ADNS servers. to shed light on this trend, we conduct a large-scale measurement to investigate the ADNS deployment patterns of modern web services and examine the characteristics of different deployment styles, such as perfor- mance, life-cycle of servers, and availability. Furthermore, we specifically focus on the DNS deployment for subdomains hosted in IaaS clouds. Then, we examine a pervasive misuse of DNS names and explore a straightforward solution to mitigate the performance penalty in DNS cache. DNS cache plays a critical role in domain name resolution, providing (1) high scalability at Root and Top-level- domain nameservers with reduced workloads and (2) low response latency to clients when the resource records of the queried domains are cached. However, the pervasive misuses of domain names, e.g., the domain names of “one-time-use” pattern, have negative impact on the effectiveness of DNS caching as the cache has been filled with those entries that are highly unlikely to be retrieved. By leveraging the domain name based features that are explicitly available from a domain name itself, we propose simple policies for improving DNS cache performance and validate their efficacy using real traces. Finally, we investigate the security implications of a fundamental vulnerability in DNS- based CDNs. The success of CDNs relies on the mapping system that leverages the dynamically generated DNS records to distribute a client’s request to a proximal server for achieving optimal content delivery. However, the mapping system is vulnerable to malicious hijacks, as it is very difficult to provide pre-computed DNSSEC signatures for dynamically generated records in CDNs. We illustrate that an adversary can deliberately tamper with the resolvers to hijack CDN’s redirection by injecting crafted but legitimate mappings between end-users and edge servers, while remaining undetectable by exist- ing security practices, which can cause serious threats that nullify the benefits offered by CDNs, such as proximal access, load balancing, and DoS protection. We further demonstrate that DNSSEC is ineffective to address this problem, even with the newly adopted ECDSA that is capable of achieving live signing for dynamically generated DNS records. We then discuss countermeasures against this redirection hijacking

Molecular evolution of candidate male reproductive genes in the brown algal model Ectocarpus

Background: Evolutionary studies of genes that mediate recognition between sperm and egg contribute to our understanding of reproductive isolation and speciation. Surface receptors involved in fertilization are targets of sexual selection, reinforcement, and other evolutionary forces including positive selection. This observation was made across different lineages of the eukaryotic tree from land plants to mammals, and is particularly evident in free-spawning animals. Here we use the brown algal model species Ectocarpus (Phaeophyceae) to investigate the evolution of candidate gamete recognition proteins in a distant major phylogenetic group of eukaryotes. Results: Male gamete specific genes were identified by comparing transcriptome data covering different stages of the Ectocarpus life cycle and screened for characteristics expected from gamete recognition receptors. Selected genes were sequenced in a representative number of strains from distant geographical locations and varying stages of reproductive isolation, to search for signatures of adaptive evolution. One of the genes (Esi0130_0068) showed evidence of selective pressure. Interestingly, that gene displayed domain similarities to the receptor for egg jelly (REJ) protein involved in sperm-egg recognition in sea urchins. Conclusions: We have identified a male gamete specific gene with similarity to known gamete recognition receptors and signatures of adaptation. Altogether, this gene could contribute to gamete interaction during reproduction as well as reproductive isolation in Ectocarpus and is therefore a good candidate for further functional evaluation

Novel 5-oxo-hexahydroquinoline derivatives: design, synthesis, in vitro P-glycoprotein-mediated multidrug resistance reversal profile and molecular dynamics simulation study

Overexpression of the efflux pump P-glycoprotein (P-gp) is one of the important mechanisms of multidrug resistance (MDR) in many tumor cells. In this study, 26 novel 5-oxo-hexahydroquinoline derivatives containing different nitrophenyl moieties at C-4 and various carboxamide substituents at C-3 were designed, synthesized and evaluated for their ability to inhibit P-gp by measuring the amount of rhodamine 123 (Rh123) accumulation in uterine sarcoma cells that overexpress P-gp (MES-SA/Dx5) using flow cytometry. The effect of compounds with highest MDR reversal activities was further evaluated by measuring the alterations of MES-SA/Dx5 cells' sensitivity to doxorubicin (DXR) using MTT assay. The results of both biological assays indicated that compounds bearing 2-nitrophenyl at C-4 position and compounds with 4-chlorophenyl carboxamide at C-3 demonstrated the highest activities in resistant cells, while they were devoid of any effect in parental nonresistant MES-SA cells. One of the active derivatives, 5c, significantly increased intracellular Rh123 at 100 mu M, and it also significantly reduced the IC50 of DXR by 70.1% and 88.7% at 10 and 25 mu M, respectively, in MES-SA/Dx5 cells. The toxicity of synthesized compounds against HEK293 as a noncancer cell line was also investigated. All tested derivatives except for 2c compound showed no cytotoxicity. A molecular dynamics simulation study was also performed to investigate the possible binding site of 5c in complex with human P-gp, which showed that this compound formed 11 average H-bonds with Ser909, Thr911, Arg547, Arg543 and Ser474 residues of P-gp. A good agreement was found between the results of the computational and experimental studies. The findings of this study show that some 5-oxo-hexahydroquinoline derivatives could serve as promising candidates for the discovery of new agents for P-gp-mediated MDR reversal