Use of Molecular Modeling to Study Spermatogenesis: An Overview Using Proteins in Sertoli Cells.

Computational structure prediction and analysis helps in understanding the structure and function of varied proteins, which otherwise becomes implausible to understand by experimental procedures. Computational techniques prove to be instrumental in understanding the molecular mechanisms that underlies physiological processes and thereby also assist in identification of potent inhibitors. Spermatogenesis, being an important cellular process that decides the fate of the progeny, holds numerous molecular interaction data, which when identified and visualized with computational structural insights, might yield a cohesive and clear-cut perception to the functionality of several proteins involved. The present chapter deals with a few selected applications of computational structure prediction towards understanding the structure of proteins and highlights how these insights are useful in providing a better understanding of different processes in spermatogenesis.

Targeting Kinase Interaction Networks: A New Paradigm in PPI Based Design of Kinase Inhibitors.

BACKGROUND: Kinases are key modulators in regulating diverse range of cellular activities and are an essential part of the protein-protein interactome. Understanding the interaction of kinases with different substrates and other proteins is vital to decode the cell signaling machinery as well as causative mechanism for disease onset and progression. OBJECTIVE: The objective of this review is to present all studies on the structure and function of few important kinases and highlight the protein-protein interaction (PPI) mechanism of kinases and the kinase specific interactome databases and how such studies could be utilized to develop anticancer drugs. METHODS: The article is a review of the detailed description of the various domains in kinases that are involved in protein-protein interactions and specific inhibitors developed targeting these PPI domains. RESULTS: The review has surfaced in depth the interacting domains in key kinases and their features and the roles of PPI in the human kinome and the various signaling cascades that are involved in certain types of cancer. CONCLUSION: The insight availed into the mechanism of existing peptide inhibitors and peptidomimetics against kinases will pave way for the design and generation of domain specific peptide inhibitors with better productivity and efficiency and the various software and servers available can be of great use for the identification and analysis of protein-protein interactions.

Computational Methods Involved in Evaluating the Toxicity of the Reproductive Toxicants in Sertoli Cell.

The Sertoli cell, the somatic component of seminiferous tubule, provides nutritional support and immunological protection and supports overall growth and division of germ cells. Cytoskeletons, junction proteins, and kinases in Sertoli cells are prime targets for reproductive toxicants and other environmental contaminants. Among the varied targets, the kinases that are crucial for regulating varied activities in spermatogenesis such as assembly/disassembly of blood-testis barrier and apical ES and those that are involved in conferring polarity are highly targeted. In an attempt to study the effect of toxicants on these kinases, the present chapter deals with computational methodology concerning their three-dimensional structure prediction, identification of inhibitors, and understanding of conformational changes induced by these inhibitors.

Effect of environmental contaminants on spermatogenesis.

Indiscriminate use of synthetic chemical compounds and the unregulated presence of heavy metals threatens the integral reproducibility of mankind and other living organisms. The toxicity of these compounds far outweighs the usefulness of these compounds. Male reproductive health is linked to the process of spermatogenesis and there is a general consensus that males are more sensitive to these environmental contaminants and so significantly affected when compared to their female counterparts. The review discusses the various toxic contaminants polluting the environment and the effect of these compounds on spermatogenesis and its relevance on male infertility in humans. It provides a detailed report on the chemical nature of few selected reprotoxicants like estrogen analogues, phthalates, dioxins, heavy metals and their action mechanism on various cellular targets that play a role in spermatogenesis with special highlights at the genetic and molecular levels. Understanding the toxicity of these compounds serves a dual purpose; to develop counter measures to protect ourselves from cellular damage and to use these compounds as a model to better understand the intricate process of spermatogenesis. The review would also help researchers formulate stringent regulations and usage restrictions in the synthesis of new compounds.

Structural insights into tumor-specific chaperoning activity of gamma synuclein in protecting estrogen receptor alpha 36 and its role in tamoxifen resistance in breast cancer.

Gamma synuclein (γSyn), a tumor-specific molecular chaperone, protects Hsp90 client proteins like ERα36 and stimulates rapid membrane-initiated estrogen signalling in breast cancer cells. However, the structural perspectives of this tumor-specific chaperone function of γSyn remains unclear. Hence, in this present work, we studied the conformational dynamics of ERα36 in the absence and presence of Hsp90 and γSyn. Results indicate that in a chaperone-free state, ERα36 undergoes an inter-domain movement and exposes the hydrophobic patch of residues that are responsible for binding with ubiquitin. However, independent of Hsp90, γSyn, by establishing transient interactions, prevents interdomain movement, unveils the co-activator binding groove, masks the ubiquitin-binding residues and maintains 'open' pocket conformation of LBD. By doing so, γSyn effectively protects ERα36 from degradation and maintains its functional state like Hsp90 based chaperoning machinery but independent of ATP. Our studies also show that the γSyn protected conformation of ERα36 can effectively bind with both estradiol (E2) and 4-hydroxy tamoxifen (4-OHT). Although they exhibit unique binding modes, they maintained the functionally active conformation of ERα36. Interestingly, the molecular dynamics simulation studies showed that 4-OHT, like γSyn, prevented the interdomain movements, primes the co-activator binding groove of ERα36 for complexation with downstream signalling proteins and this mechanism explains its agonist activity and associated anti-estrogen resistance observed in the presence of ERα36. The observed differences in the chaperoning mechanism of γSyn sheds light on its selectivity over Hsp90 in cancer cells, for promoting rapid protection of crucial oncogenic proteins. Based on our findings, we speculate that the compounds, which can hamper association of γSyn with ERα36 and/or can arrest ERα36 in an ubiquitin binding state, would be promising alternatives for treating ERα36 expressed breast carcinomas.

Structural perspective of ARHI mediated inhibition of STAT3 signaling: an insight into the inactive to active transition of ARHI and its interaction with STAT3 and importinß.

ARHI, a putative tumor suppressor protein with unique 32 amino acid extension in the N-terminal region, differs from oncogenes Ras and Rap, negatively regulates STAT3 signaling and inhibits the migration of ovarian cancer cells. ARHI associates directly with STAT3, also forms complex with importinß, and prevents formation of RanGTPase-importinß complex, which is essential for transporting STAT3 into the nucleus. Hence, the structural aspects pertaining to ARHI mediated inhibition of STAT3 translocation can provide hints on the regulation of STAT3 signaling mechanism. Accordingly, in the present study, the structure of ARHI was predicted and its transition from inactive to active state studied using MD simulations and free energy landscape analysis. The transition of ARHI is marked by the movement of switch I region towards γ-phosphate of GTP, in addition, the hydrophobic interaction between N-terminal helix and switch II region of ARHI accounts for its low intrinsic GTPase activity. Further, the protein-protein interaction studies reveal that the residues of N-terminal helix, effector domain, P-loop and G box motif of ARHI actively form polar and non-polar interaction with NTD of STAT3 and make them compact thereby rendering STAT3 inaccessible for Ran-importinß mediated translocation. On the other hand, ARHI competes with RanGTPase and interacts with importinß via basic-acidic patch interaction, which leads to inhibition of STAT3 translocation. The interacting residues involved for this structural mechanism would be instrumental in designing inhibitors for STAT3, which mimics ARHI thereby leading to the suppression of cancer cell growth.

Molecular dynamic simulations of the tubulin-human gamma synuclein complex: structural insight into the regulatory mechanism involved in inducing resistance against Taxol.

Members of the synuclein family (α, ß and γ synucleins) are intrinsically disordered in nature and play a crucial role in the progression of various neurodegenerative disorders and cancers. The association of γSyn with both BubR1 as well as microtubule subunits renders resistance against various anti-cancer drugs. However, the structural aspects underlying drug resistance have not been explored. In this study, the mechanism involved in the association between γSyn and microtubule subunits (αßTub) was investigated and the results reveal a strong interaction between γSyn and the tail regions of αßTub. Complexation of γSyn induces conformational rearrangements in the nucleotide binding loops (NBL), interdomain and tail regions of both α and ßTub. Moreover, in ßTub, the massive displacement observed in M and S loops significantly alters the binding site of microtubule targeting drugs like Taxol. The resulting weak association between Taxol and ßTub of the γSyn-αßTub complex was confirmed by molecular dynamic simulation studies. In addition, the effect of Taxol on NBL, M and S loops of αßTub, is reversed in the presence of γSyn. These results clearly indicate that the presence of γSyn annulled the allosteric regulation imposed by Taxol on the αßTub complex as well as preventing the binding of microtubule targeting drugs, which eventually leads to the development of resistance against these drugs in cancer cells.

Structural insights into interacting mechanism of ID1 protein with an antagonist ID1/3-PA7 and agonist ETS-1 in treatment of ovarian cancer: molecular docking and dynamics studies.

Among the many abnormally expressed proteins in ovarian cancer, the prominent cancer in women, ID1 (inhibitors of DNA binding protein 1) is a potential one among other several targets. Interaction of ID1 with ETS-1 (transcriptional activator of p16(INK4a)) suppresses the transcription of p16(INK4a) and causes abnormal cell proliferation. A peptide aptamer (ID1/3-PA7) has been designed to prevent this interaction and thereby leading to the transcription of p16(INK4a). However, the structural basis behind the molecular interaction of ID1 with ETS-1 (agonist) and ID1/3-PA7 (antagonist) is poorly understood. In order to understand this structural recognition and their interaction mechanism, in silico methods were used. From this interaction analysis, the residues of ETS-1 involved in interaction with the p16(INK4a) promoter were found to be targeted by ID1. Subsequently, ETS-1 binding residues of ID1 were found to be targeted by its aptamer- ID1/3-PA7. These results suggest that both ETS-1 and ID1/3-PA7 binds at the same region harbored by the residues-H97, D100, R103, D104, L107, A144, C145, D149, D150 and C154 of ID1. All these observations correlate with the experimental reports, suggesting that the identified residues might play a crucial role in promulgating the oncogenic effects of ID1. In silico alanine scanning mutagenesis also confirms the role of identified hot spot residues in p16(INK4a) regulation. Finally, the molecular dynamic simulation studies reveal the prolonged stability of the aforementioned interacting complexes. The obtained results throw light on the structure and residues of ID1 involved in transcriptional regulation of p16(INK4a).

Molecular dynamics simulation of the Staphylococcus aureus YsxC protein: molecular insights into ribosome assembly and allosteric inhibition of the protein.

YsxC from Staphylococcus aureus is a member of the GTPase protein family, and is involved in the ribosomal assembly and stability of this microorganism through its interactions with the L17, S2 and S10 ribosomal proteins. Inhibition of its interactions with L17, S2, S10 and the ß' subunit of RNA polymerase influences ribosomal assembly, which may affect the growth of the microorganism. This makes YsxC a novel target for the design of inhibitors to treat the disease caused by S. aureus. Understanding the interaction mechanism between YsxC and its partners would aid in the identification of potential catalytic residues, which could then be targeted to inhibit its function. Accordingly, in the present study, an in silico analysis of the interactions between YsxC and L17, S2 and S10 was performed, and the potential residues involved in these interactions were identified. Based on the simulation results, a possible mechanism for the interactions between these proteins was also proposed. Finally, six ligands from among a library of 81,000 chemical molecules were found to interact with parts of the G2 and switch II regions of the YsxC protein. Moreover, their interactions with the YsxC protein were observed to provoke changes at its GTP-binding site, which suggests that the binding of these ligands leads to a reduction in GTPase activity, and they were also found to affect the interactions of YsxC with its partners. This observation indicates that the proposed interacting site of YsxC may act as an allosteric site, and disrupting interactions at this site might lead to novel allosteric inhibition of the YsxC protein.