Yeast centromere DNA (CEN) affinity column chromatography has been used to purify several putative and kinetochore proteins from yeast chromatin extracts. The single gene (CBF5) specifying one of the major low-affinity centromere-binding (p64'/CBF5p) cloned shown be essential for viability Saccharomyces cerevisiae. CBF5 specifies a 55-kDa highly charged protein that contains repeating KKD/E sequence domain near C terminus, similar known microtubule-binding domains in microtubule-associated...

A set of high-affinity, high-specificity posttranslational modification (PTM) enrichment tools was developed to generate an unbiased snapshot four key PTM profiles (tyrosine phosphorylation, acetylation, ubiquitination, and SUMOylation 2/3) for the clinically important protein programmed cell death ligand 1 (PD-L1). The results showed that epidermal growth factor (EGF) treatment induced tyrosine ubiquitination PD-L1. Further characterization EGF-induced PD-L1 revealed a significant increase...

We have used in vitro motility assays to investigate the mechanism of kinetochore function budding yeast Saccharomyces cerevisiae. Functional centromeric DNA plus a tripartite centromere binding protein complex, CBF3, was found be necessary but not sufficient for activity. A fourth required component identified as motor Kar3p, previously reported kinesin known involved karyogamy and mitosis. Our data support genetic evidence suggesting that Kar3p is kinetochore-associated imply CBF3 plays...

Yeast centromere DNA (CEN) affinity column chromatography has been used to purify several putative and kinetochore proteins from yeast chromatin extracts. The single gene (CBF5) specifying one of the major low-affinity centromere-binding (p64'/CBF5p) cloned shown be essential for viability Saccharomyces cerevisiae. CBF5 specifies a 55-kDa highly charged protein that contains repeating KKD/E sequence domain near C terminus, similar known microtubule-binding domains in microtubule-associated...

The Xenopus laevis alpha-tubulin gene X alpha T14, which is highly expressed during oogenesis, exhibits accurate and efficient transcription initiation when microinjected into X. oocytes. However, we found previously in nuclease protection assays of transcripts from injected T14 that many protected fragments were shorter than expected could be produced. We show here by exonuclease VII mapping, Northern (RNA) blotting, gel fractionation RNA these caused truncated share the same sites as...

Post-translational modification (PTM) crosstalk is recognized as a major cell-regulatory mechanism, and studies of several proteins have validated the premise that PTMs work in concert. Previous by our group investigated potential PTM on EGFR-Ras-c-Fos axis utilizing comprehensive set reagents termed Signal-Seeker toolkits. In this study, these tools were used to investigate occurs acetylated mitochondrial response stress-inducing agent hydrogen peroxide (H2O2). Mitochondrial protein...

It is presently accepted that the mechanism of action for all anti-tumor tubulin ligands involves perturbation microtubule dynamics during G2/M phase cell division and subsequent entry into apoptosis 1]. In this report, we challenge established dogma by describing a unique caused novel series ligands, halogenated derivatives acetamido benzoyl ethyl ester. We have developed suicide ligand tubulin, which covalently attaches to target shows potent cancericidal activity in tissue culture assays...

We have identified in an intron of X. laevls α-tubulin gene a member novel family short (226 – 431 bp) interspersed repetitive elements. isolated other members this family, which we term Ocr, from ovary cDNA and genome libraries another two the published sequences H1B histone cluster actin intron. The termini Ocr elements are formed by 19 bp inverted repeat that has clear sequence homologies to those certain large transposable elements, such as 1723 (Xenopus) Ac (maize). However, do not...

Identification of a novel post-translational modification (PTM) for target protein, defining its physiologic role and studying potential cross-talk with other PTMs is challenging process. A set highly sensitive tools termed as Signal-Seeker kits was developed, which enables rapid simple detection on any protein. The methodology these utilizes affinity purification modified proteins from cell or tissue lysate, immunoblot analysis. These utilize single lysis system that effective at...

We have studied the expression of XαT14, a member α-tubulin multigene family in Xenopus laevis. Small amounts XαT14 RNA are detectable range cell types, but much higher levels present ovary and tissue culture cells. In oocytes transcripts accumulate during early vitellogenesis their level declines more advanced stages. Faithful efficient initiation transcription occurred on cloned injected into even at low template levels. examined amount transcript produced by various deletion mutants...

It is now well-appreciated that post-translational modifications (PTMs) play an integral role in regulating a protein's structure and function, which may be essential for given both physiologically pathologically.Enrichment of PTMs often necessary when investigating the PTM status target protein, because are transient relatively low abundance.Many pitfalls encountered enriching such as buffer incompatibility, protein antibody not IP-compatible, loss signal, others.The degree difficulty...

Lysine acetylation is an important regulatory post-translational modification (PTM) that occurs sub-stoichiometrically, often representing less than 1% of the target protein. This makes studying endogenous protein extremely challenging. Recent reports suggest several modifications (PTMs), including lysine acetylation, play a major role in regulation programmed cell death-ligand 1 (PD-L1), clinically target. An enrichment step necessary to enable identification acetylated species by either...

It is now well-appreciated that post-translational modifications (PTMs) play an integral role in regulating a protein's structure and function, which may be essential for given both physiologically pathologically. Enrichment of PTMs often necessary when investigating the PTM status target protein, because are transient relatively low abundance. Many pitfalls encountered enriching such as buffer incompatibility, protein antibody not IP-compatible, loss signal, others. The degree difficulty...