Enzyme Kinetics Features of the Representative Engineered Recombinants of Chondroitinase ABC I.

Chondroitinase ABC I (cABC I) from Proteus vulgaris is an important enzyme in medicinal biotechnology due to its ability to help axon regeneration after spinal cord injury. Its practical application involves solving several problems at the molecular and cellular levels. Structurally, most residues at the C-terminal domain of cABC I are arranged as organized strands, and only a small fraction of residues have helical conformation. The structural and functional features of modified residues on two specific helix fragments have previously been reported. The single mutant M889K has been combined with L679S and L679D mutants to make enzyme variants containing simultaneously modified helix. Here, the pH stability and temperature-based analysis of the transition state structure for the catalysis reaction were investigated. We found that double mutant L679D/M889K is the better choice to use in physiological conditions due to its higher pH stability at physiological pH as well as its different optimum temperature as compared with the (wild-type) WT protein. According to Arrhenius's analysis, the values of the Gibbs free energy of the transition state (∆G#) are not changed upon mutation. However, the relative contribution and absolute values of the enthalpy and entropy change to the total value of ∆G#, varied between the WT and mutants.

Two Novel Tri-Aryl Derivatives Attenuate the Invasion-Promoting Effects of Stromal Mesenchymal Stem Cells on Breast Cancer.

BACKGROUND: The concept of Epithelial-Mesenchymal Transition (EMT) to promote carcinoma progression has been recognized as a venue for research on novel anticancer drugs. Triaryl template-based structures are one of the pivotal structural features found in a number of compounds with a wide variety of biological properties including anti-breast cancer. Among the various factors triggering EMT program, cyclooxygenase-2 (COX-2), NF-κB as well as the transforming growth factor-beta (TGF-ß) have been widely investigated. OBJECTIVE: Here, we aim to investigate the effect of two novel compounds A and B possessing triaryl structures, which interact with both COX-2 and TGF-ß active sites and suppress NF-κB activation, on EMT in a co-culture system with breast cancer and stromal cells. METHODS: MDA-MB-231 and bone-marrow mesenchymal stem (BM-MS) cells were co-cultured in a trans-well plate. Migration, matrigel-based invasion and colony formation in soft agar assays along with Real- time PCR and Western blot analysis were performed to examine the effect of compounds A and B on the invasive properties of MDA-MB-231 cells after 72 hours of co-culturing with BM-MSCs. In addition, TGF-beta interaction was investigated by Localized Surface Plasmon Resonance (LSPR). RESULTS: BM-MSCs enhanced migration, invasion and anchorage-independent growth of the co-cultured MDAMB- 231 cells. A reduction in E-cadherin level concomitant with an increase in vimentin and N-cadherin levels following the co-culture implied EMT as the underlying process. Compounds A and B inhibited invasion and anchorage-independent growth of breast cancer cells co-cultured with BM-MSCs at 10µM. The observed inhibitory effects along with an increase in E-cadherin and a reduction in vimentin and ZEB2 levels suggest that the anti-invasive properties of compounds A and B might proceed through the blockade of stromal cell-induced EMT, mediated by their interaction with TGF-beta. CONCLUSION: These findings introduce compounds A and B as novel promising agents, which prevent EMT in invasive breast cancer cells.

Investigating the structural and functional features of representative recombinants of chondroitinase ABC I.

Chondroitin Sulfate Proteoglycans (CSPGs) are the main inhibitors for axon regeneration after damaging of Central Nervous System (CNS). Chondroitinase ABC I (cABC I) can degrade CSPGs by removing chondroitin and dermatan sulfate side chains from proteoglycans. Hence, it may be considered as an attractive candidate in biomedicine. For practical applications of this enzyme, increasing the effective circulating level and reducing the number and volume of injections for patients is one of the main concerns which is directly related to conformational stability and catalytic efficiency of the enzyme. Structural examination of C-terminal domain of cABC I reveals that there are a few numbers of residues in helical conformation which are positioned at the context of a cohesive structural organization of ß-strands. In line with our previous studies on C-terminal domain of cABC I and regarding the residues in α-helix conformation; we designed and constructs some representative mutants including M889K, M889L, L679D/M889K and L679S/M889K. According to structural and functional characterization of protein variants and regarding the wide range of variability in determining parameters for ß-sheet conformation, we proposed a model in which the structural integrity of ß-strands at C-terminal domain can be manipulated and directed toward a new patterns of organization, some of them may have positive effects on the structural and functional features of the enzyme. Using this strategy it may be possible to improve functional and structural features of the enzyme by engineering the intra-molecular interactions in positions far from the active site of the enzyme.

Corrigendum: The Associations of Dyadic Coping and Relationship Satisfaction Vary between and within Nations: A 35-Nation Study.

[This corrects the article on p. 1106 in vol. 7, PMID: 27551269.].

The Associations of Dyadic Coping and Relationship Satisfaction Vary between and within Nations: A 35-Nation Study.

OBJECTIVE: Theories about how couples help each other to cope with stress, such as the systemic transactional model of dyadic coping, suggest that the cultural context in which couples live influences how their coping behavior affects their relationship satisfaction. In contrast to the theoretical assumptions, a recent meta-analysis provides evidence that neither culture, nor gender, influences the association between dyadic coping and relationship satisfaction, at least based on their samples of couples living in North America and West Europe. Thus, it is an open questions whether the theoretical assumptions of cultural influences are false or whether cultural influences on couple behavior just occur in cultures outside of the Western world. METHOD: In order to examine the cultural influence, using a sample of married individuals (N = 7973) from 35 nations, we used multilevel modeling to test whether the positive association between dyadic coping and relationship satisfaction varies across nations and whether gender might moderate the association. RESULTS: RESULTS reveal that the association between dyadic coping and relationship satisfaction varies between nations. In addition, results show that in some nations the association is higher for men and in other nations it is higher for women. CONCLUSIONS: Cultural and gender differences across the globe influence how couples' coping behavior affects relationship outcomes. This crucial finding indicates that couple relationship education programs and interventions need to be culturally adapted, as skill trainings such as dyadic coping lead to differential effects on relationship satisfaction based on the culture in which couples live.