Epidermal growth factor receptors: function modulation by phosphorylation and glycosylation interplay

Post-translational modifications (PTMs) of proteins induce structural and functional changes that are most often transitory and difficult to follow and investigate invivo. Insilico prediction procedures for PTMs are very valuable to foresee and define such transitory changes responsible for the multifunctionality of proteins. Epidermal growth factor receptor (EGFR) is such a multifunctional transmembrane protein with intrinsic tyrosine kinase activity that is regulated primarily by ligand-stimulated transphosphorylation of dimerized receptors. In human EGFR, potential phosphorylation sites on Ser, Thr and Tyr residues including five autophosphorylation sites on Tyr were investigated using insilico procedures. In addition to phosphorylation, O-GlcNAc modifications and interplay between these two modifications was also predicted. The interplay of phosphorylation and O-GlcNAc modification on same or neighboring Ser/Thr residues is termed as Yin Yang hypothesis and the interplay sites are named as Yin Yang sites. Amongst these modification sites, one residue is localized in the juxtamembrane (Thr 654) and two are found in the catalytic domain (Ser 1046/1047) of the EGFR. We propose that, when EGFR is O-GlcNAc modified on Thr 654, EGFR may be transferred from early to late endosomes, whereas when EGFR is O-GlcNAc modified on Ser 1046/1047 desensitization of the receptor may be prevented. These findings suggest a complex interplay between phosphorylation and O-GlcNAc modification resulting in modulation of EGFR's functionalit

Oct-2 DNA binding transcription factor: functional consequences of phosphorylation and glycosylation

Phosphorylation and O-GlcNAc modification often induce conformational changes and allow the protein to specifically interact with other proteins. Interplay of phosphorylation and O-GlcNAc modification at the same conserved site may result in the protein undergoing functional switches. We describe that at conserved Ser/Thr residues of human Oct-2, alternative phosphorylation and O-GlcNAc modification (Yin Yang sites) can be predicted by the YinOYang1.2 method. We propose here that alternative phosphorylation and O-GlcNAc modification at Ser191 in the N-terminal region, Ser271 and 274 in the linker region of two POU sub-domains and Thr301 and Ser323 in the POUh subdomain are involved in the differential binding behavior of Oct-2 to the octamer DNA motif. This implies that phosphorylation or O-GlcNAc modification of the same amino acid may result in a different binding capacity of the modified protein. In the C-terminal domain, Ser371, 389 and 394 are additional Yin Yang sites that could be involved in the modulation of Oct-2 binding propertie

Terminal Galactose as Cancer Recognition Marker: Computing Analysis With Implications of Vicinal Sugars, Linkage and Anomery

Abstract.-Carbohydrate structures on proteins play vital role as recognition markers in several diseases including cancer. The carbohydrate chains are dramatically altered in cancer cells compared to normal cells, both in structure and quantity. The mucin O-glycans show several cancer associated structures like T-(Gal-GalNAc-Ser/Thr) and Tn-antigen (GalNAc-Ser/Thr). Terminal galactose (Gal) in mucin type O-glycan oligosaccharide structures is known to act as a recognition marker for several cancer-associated lectins like galectin and mistletoe lectins. This study describes the role of terminal Gal, inclusive of its anomery, linkage and that of sugar residues vicinal to terminal Gal in oligosaccharide structures of glycoproteins, as an epitope or as a recognition marker in cancer

Somatic Cell Cycle Regulation By Histone H3 Modifications: Action of OGT and Kinases

Abstract.-Histone H3 is amongst the most evolutionarily conserved proteins, and is located along with histone 2A, 2B and 4 in the core of the nucleosome. The N-terminal tails of the histone protrude the chromatin structure and become accessible to various enzymes for post translational modifications (PTMs). Phosphorylation of H3 has been found to have an impact on progression of the cell cycle, especially during mitosis. Another equally abundant PTM is the glycosylation at serine/threonine by O-GlcNAc (O-linked glycosylation) that occurs on the same or neighboring Ser or Thr residues, which also are accessible to kinases (Yin Yang sites). O-GlcNAc is added by O-GlcNAc transferase (OGT), and is found exclusively in the nucleus or cytoplasm of the cell. By using computational methods like Netphos 2.0 and Yinoyang 1.2 we found that OGT, Aurora B kinase and OGT, Death-associated protein (DAP)-like kinase, work together in a Yin Yang way, and thereby control specific checkpoints during mitosis. Bioinformatics tool, thus, are very helpful to elucidate the function of the protein by predicting the PTMs in proteins

Galactose: A Specifically Recognized, Terminal Carbohydrate Moiety in Biological Processes

Glycoproteins and glycolipids carrying diverse oligosaccharide structures are involved in countless molecular interactions in physiologic and pathologic situations. Defining the specific carbohydrate moieties expressed in a particular set of molecules is a challenging task that could eventually explain how glycoproteins and glycolipids contribute to the physiology of normal cells and how their alterations could lead to pathologic states. A simple example is the ABO blood group system: in individuals with blood group B, the marker is defined by its terminal linked galactose, and substitution of its hydroxyl group at C2 by an N-acetyl group results in the formation of N-acetylgalactosamine, the blood group A marker. This review focuses on the importance of terminal linked galactose and its derivatives in different normal and pathological conditions. The involvement of various sugars residues sub-terminal to galactose and its derivatives was also evaluated on the basis of the galactosylation data taken from different publicly available carbohydrate databases. We conclude that those sugars penultimate to galactose, with their different types of linkages and anomery, contribute to the structure and functions of carbohydrate moieties with a terminal galactose

Role of sialic acid and sulfate groups in cervical mucus physiological functions: study of Macaca radiata glycoproteins

The influence of charged groups in glycoproteins was investigated to assess their effect on the physiological functions of bonnet monkey cervical mucus. The macromolecular glycoproteins from peri-ovulatory, midcycle phase cervical mucus were treated with Pronase, trypsin and chymotrypsin and the enzyme-resistant glycoproteins purified by gel filtration on Sepharose 4B and a high molecular weight component containing carbohydrates, proteins and sulfate groups was recovered in high yield. This material still reacted with an antiserum directed against purified midcycle glycoprotein but not against another antiserum directed against luteal phase purified glycoproteins. Upon treatment with Pronase, trypsin and chymotrypsin, asialoglycoproteins and desulfated asialoglycoproteins released fragments of low molecular sizes, none of which reacted with the anti-midcycle glycoprotein antiserum. Cervical mucus collected from the estrogenic phase displayed a morphology supporting sperm migration, and this mucus retains the same morphology and reacts with the anti-midcycle glycoprotein antiserum following mild treatment with sialidase and subsequently with Pronase. These results imply that charged carbohydrate groups help maintain the structural and functional integrity of the mucus glycoprotein in its biological environment

Oct-2 DNA binding transcription factor: functional consequences of phosphorylation and glycosylation

Phosphorylation and O-GlcNAc modification often induce conformational changes and allow the protein to specifically interact with other proteins. Interplay of phosphorylation and O-GlcNAc modification at the same conserved site may result in the protein undergoing functional switches. We describe that at conserved Ser/Thr residues of human Oct-2, alternative phosphorylation and O-GlcNAc modification (Yin Yang sites) can be predicted by the YinOYang1.2 method. We propose here that alternative phosphorylation and O-GlcNAc modification at Ser191 in the N-terminal region, Ser271 and 274 in the linker region of two POU sub-domains and Thr301 and Ser323 in the POUh subdomain are involved in the differential binding behavior of Oct-2 to the octamer DNA motif. This implies that phosphorylation or O-GlcNAc modification of the same amino acid may result in a different binding capacity of the modified protein. In the C-terminal domain, Ser371, 389 and 394 are additional Yin Yang sites that could be involved in the modulation of Oct-2 binding properties

Effect on the Ras/Raf signaling pathway of post-translational modifications of neurofibromin: in silico study of protein modification responsible for regulatory pathways

Mapping and chemical characterization of post-translational modifications (PTMs) in proteins are critical to understand the regulatory mechanisms involving modified proteins and their role in disease. Neurofibromatosis type 1 (NF-1) is an autosomal dominantly inherited disorder, where NF1 mutations usually result in a reduced level of the tumor suppressor protein, neurofibromin (NF). NF is a multifunctional cytoplasmic protein that regulates microtubule dynamics and participates in several signaling pathways, particularly the RAS signaling pathway. NF is a Ras GTPase-activating protein (GAP) that prevents oncogenesis by converting GTP-Ras to GDP-Ras. This function of NF is regulated by phosphorylation. Interplay of phosphorylation with O-GlcNAc modification on the same or vicinal Ser/Thr residues, the Yin Yang sites, is well known in cytoplasmic and nuclear proteins. The dynamic aspects of PTMs and their interplay being difficult to follow in vivo, we undertook this in silico work to predict and define the possible role of Yin Yang sites in NF-1. Interplay of phosphorylation and O-GlcNAc modification is proposed as a mechanism controlling the Ras signaling pathway