Biology subjects

Lei, M.

Publications and source records attributed to Lei, M..

4 recordsLinked to original sources

bioRxiv2018

Crystal structure of the MBD domain of MBD3 in complex with methylated CG DNA

MBD3 is a core subunit of the Mi-2/NuRD complex, and has been previously reported to lack methyl-CpG binding ability. However, recent reports show that MBD3 recognizes both mCG and hmCG DNA with a preference for hmCG, and is required for the normal expression of hmCG marked genes in ES cells. Nevertheless, it is not clear how MBD3 recognizes the methylated DNA. In this study, we carried out structural analysis coupled with isothermal titration calorimetry (ITC) binding assay and mutagenesis studies to address the structural basis for the mCG DNA binding ability of the MBD3 MBD domain. We found that the MBD3 MBD domain prefers binding mCG over hmCG through the conserved arginine fingers, and this MBD domain as well as other mCG binding MBD domains can recognize the mCG duplex without orientation selectivity. Furthermore, we found that the tyrosine-to-phenylalanine substitution at Phe34 of MBD3 is responsible for its weaker mCG DNA binding ability compared to other mCG binding MBD domains. In summary, our study demonstrates that the MBD3 MBD domain is a mCG binder, and also illustrates its binding mechanism to the methylated CG DNA.

biochemistry↗

bioRxiv2018

Structural and histone binding studies of the chromo barrel domain of TIP60

TIP60 consists of an N-terminal chromo barrel domain (TIP60-CB) and a C-terminal acetyltransferase domain and acetylates histone and non-histone proteins within diverse cellular processes. Whereas the TIP60-CB is thought to recognize histone tails, molecular details of this interaction remain unclear. Here we attempted a quantitative analysis of the interaction between the TIP60-CB and histone peptides, but did not observe any binding through either fluorescence polarization or isothermal titration calorimetry. We solved a crystal structure of the TIP60-CB alone. Analysis of the crystal structure demonstrates a putative peptide binding site that may be occluded by the basic side chain of a residue in a unique {beta} hairpin between the two N-terminal strands of the {beta} barrel.

biochemistry↗

bioRxiv2018

Crystal structure of chromo barrel domain of RBBP1

RBBP1 is a retinoblastoma protein (pRb) binding protein acting as a repressor of gene transcription. RBBP1 is a multidomain protein including a chromo barrel domain, and its chromo barrel domain has been reported to recognize histone H4K20me3 weakly, and this binding is enhanced by the simultaneous binding of DNA. However, the molecular basis of this DNA-mediated histone binding by the chromo barrel domain of RBBP1 is unclear. Here we attempted to co-crystallize the chromo barrel domain of RBBP1 with either a histone H4K20me3 peptide alone or with both a histone H4K20me3 peptide and DNA, but only solved the peptide/DNA unbound crystal structure. Our structural analysis indicates that RBBP1 could interact with histone H4K20me3 similar to other histone binding chromo barrel domains, and the surface charge representation analysis of the chromo barrel domain of RBBP1 suggests that the chromo barrel domain of RBBP1 does not have a typical DNA binding surface, indicating that it might not bind to DNA. Consistently, our ITC assays also showed that DNA does not significantly enhance the histone binding ability of the chromo barrel domain of RBBP1.

biochemistry↗

bioRxiv2017

Gossypol biosynthesis in cotton revealed through organ culture, plant grafting and gene expression profiling

Gossypol plays an important role in defense mechanism of Gossypium species and the presence of gossypol also limits the utilization of cottonseeds. However, little is known about the metabolism of gossypol in cotton plant. Here, Detection on the dynamic tendency of gossypol content illustrated that at the germination stage, the main source of gossypol was cotyledon, and at the later stages, gossypol mainly came from root system. Plant grafting between cottons and sunflower proved that gossypol was mainly synthesized in the root systems of cotton plants and both of the glanded and glandless cottons had the ability of gossypol biosynthesis. Besides, the pigment glands expression was uncoupled with gossypol biosynthesis. Root tip and rootless seedling organ culture in vitro further revealed other parts of the seedlings also got the ability to synthesize gossypol except root system. Moreover, root system produced the racemic gossypol and plant synthesized the optically active gossypol. The expression profiling of key genes in the gossypol biosynthetic pathway suggested that downstream key genes had relatively high expression levels in root systems which confirmed that gossypol was mainly synthesized in the root systems. Taken together, our results helped to clarify the complex mechanism of gossypol metabolism.

synthetic biology↗