A heparin binding protein whose expression increases during differentiation of embryonal carcinoma cells to parietal endoderm cells: cDNA cloning and sequence analysis

A cDNA clone isolated from a lambda gt11 expression library of teratocarcinoma OTT6050 specifies for a glycoprotein with a molecular weight of about 44,000. The new glycoprotein was termed heparin binding protein-44 (HBP-44), since it was absorbed to a heparin-agarose column and was eluted from it by a buffer containing 1.5 M NaCl. HBP-44 mRNA was intensely expressed in PYS-2 parietal endoderm cells and in the kidney, and the RNA level increased about 10-fold during differentiation of F9 embryonal carcinoma cells to parietal endoderm cells. From the cDNA sequence, HBP-44 was concluded to be rich in charged amino acids, and large segments of the protein appeared to form alpha-helixes. The protein was considered to be anchored to the membrane by a cluster of hydrophobic amino acids present in the N-terminal region. Indeed, the N-terminal sequence of HBP-44 was homologous to asialoglycoprotein receptor, which is anchored to the membrane by the N-terminal region. Furthermore, a portion of the N-terminal region of HBP-44 was homologous to the leucine zipper domain. Except for the N-terminal region, HBP-44 had over-all homology with structural proteins such as myosin heavy chain. We propose that HBP-44 is extruded from plasma membranes and interacts with heparin and related molecules and that it is involved in the interactions of plasma membranes with basement membranes.     

Chromosomal locations of the genes for histones and a histone gene-binding protein family HBP-1 in common wheat

The chromosomal locations of the genes in common wheat that encode the five histones and five members of the HBP (histone gene-binding protein)-1 family were determined by hybridizing their cloned DNAs to genomic DNAs of nullitetrasomic and telosomic lines of common wheat, Triticum aestivum cv. Chinese Spring. The H1 and H2a genes are located on different sets of homoeologous chromosomes or chromosome arms, namely, 5A, 5B and 5D, and 2AS, 2BS and 2DS, respectively. Genes for the other histones, H2b, H3 and H4, are found in high copy number and are dispersed among a large number of chromosomes. The genes for all members of the HBP-1 family are present in small copy numbers. Those for HBP-1a(1) are located on six chromosome arms, 3BL, 5AL, 5DL, 6AL, 6BS and 7DL, whereas those for each HBP-1a(c14), 1a(17), 1b(c1), and 1b(c38) are on a single set of homoeologous chromosome arms; 4AS, 4BL, 4DL; 6AS, 6BS, 6DS; 3AL, 3BL, 3DL; and 3AS, 3BS, 3DS, respectively. The genes for histones H1 and H2a, and for all members of the HBP-1 family except HBP-1a(1) are assumed to have different phylogenetic origins. The genes for histone 2a and HBP-1a(17) are located in the RFLP maps of chromosomes 2B and 6A, respectively. Gene symbols are proposed for all genes whose chromosomal locations have been determined.     

Identification of a heparin-binding protein in bovine seminal fluid as tissue inhibitor of metalloproteinases-2

Presence or absence of three distinct bovine seminal heparin-binding proteins (21-31 kDa) recognized in sperm extracts by a monoclonal antibody, M1, is a diagnostic indicator of fertility differences among bulls producing normal semen. We recently identified a 31 kDa fertility-associated antigenin bovine seminal fluid as a unique DNase I-like protein. We now report purification and identification of a 24 kDa seminal heparin-binding protein (HBP-24) recognized by M1. N-terminal microsequence analysis of HBP-24 purified from seminal fluid yielded 20 amino acid residues that displayed 90% identity to the N-terminus of a bovine metalloproteinase inhibitor identified as tissue inhibitor of metalloproteinases-2 (TIMP-2). A single immunoreactive band migrating at 24 kDa was detected in Western blots of cauda epididymal sperm extracts following incubation with purified seminal heparin-binding proteins and subsequent washing in vitro, indicating TIMP-2 bound to sperm membranes. Expression of TIMP-2 mRNA was detected by RT-PCR in bovine bulbourethral gland, prostate, and seminal vesicles. Mobility of the 24 kDa heparin-binding protein increased under nonreducing SDS-PAGE to approximately 21 kDa, characteristic of the reported molecular mass of TIMP-2. To our knowledge, this is the first report of TIMP-2 binding to spermatozoa and of TIMP-2 mRNA expression in bovine accessory sex glands. These results corroborate previous reports regarding the site of production of heparin-binding proteins that are related to bull fertility, and suggest that TIMP-2 influences fertility of bulls, either through inhibition of metalloprotease activity in semen or via undefined activities independent of matrix metalloproteinase (MMP) inhibition.     

HIV-1 Nef protein: purification, crystallizations, and preliminary X-ray diffraction studies.

Human immunodeficiency virus Nef protein accelerates virulent progression of AIDS by its interaction with specific cellular proteins involved in cellular activation and signal transduction. Here we report the purification and crystallization of the conserved core of HIV-1LAI Nef protein in the unliganded form and in complex with the wild-type SH3 domain of the P59fyn protein-tyrosine kinase. One-dimensional NMR experiments show that full-length protein and truncated fragment corresponding to the product of HIV-1 protease cleavage have a well-folded compact tertiary structure. The ligand-free HIV-1 Nefcore protein forms cubic crystals belonging to space group P23 with unit cell dimensions of a = b = c = 86.4 A. The Nef-Fyn SH3 cocrystals belong to the space group P6(1)22 or its enantiomorph, P6(5)22, with unit cell dimensions of a = b = 108.2 A and c = 223.7 A. Both crystal forms diffract to a resolution limit of 3.0 A resolution using synchrotron radiation, and are thus suitable for X-ray structure determination.     

G Ⅱ.23基因型诺如病毒P蛋白的寡糖结合特征

诺如病毒(Noroviruses,NoVs)是导致人急性胃肠炎的最重要病原体之一,也是引起食源性疾病暴发的首要病原体。组织血型抗原(Histo-blood groups antigens,HBGAs)是NoVs的受体或宿主易感因子。已有研究表明HBGAs与NoVs的感染和流行高度相关。GⅡ.23是最近报道的NoVs新基因型。为了研究GⅡ.23与HBGAs的结合特征,表达纯化GⅡ.23基因型的P蛋白之后,通过唾液和寡糖结合实验研究其与HBGAs的结合特性,并通过同源结构模拟探索GⅡ.23 P蛋白与糖抗原潜在的对接分子机制,与已经解析的GⅡ.10的P蛋白与岩藻糖的复合物结构进行重叠。结果发现,GⅡ.23 P蛋白可以与B型唾液结合,但不结合A、O^+和O^-非分泌型唾液;P蛋白与H双糖抗原发生结合;分子模拟显示GⅡ.23 P蛋白具有与岩藻糖环结合的类似特征。本研究首次揭示了GⅡ.23 P蛋白与HBGAs受体的结合特征,为深入探索GⅡ.23基因型NoVs的进化、感染以及流行的具体机制提供了基础资料。