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L Xue

Publications and source records attributed to L Xue.

At least 19 recordsLinked to original sources
PubMed2001

Intermembrane transfer of polyethylene glycol-modified phosphatidylethanolamine as a means to reveal surface-associated binding ligands on liposomes.

In order to explore the use of exchangeable poly(ethylene glycol) (PEG)-modified diacylphosphatidylethanolamines (PE) to temporarily shield binding ligands attached to the surface of liposomes, a model reaction based on inhibition and subsequent recovery of biotinylated liposome binding to streptavidin immobilized on superparamagnetic iron oxide particles (SA magnetic particles) was developed. PEG-lipid incorporation into biotinylated liposomes decreased liposome binding to SA magnetic particles in a non-linear fashion, where as little as 0.1 mol% PEG-PE resulted in a 20% decrease in binding. Using an assay based on inhibition of binding, PEG(2000)-PE transfer from donor liposomes to biotinylated acceptor liposomes could be measured. The influence of temperature and acyl chain composition on the transfer of PEG-diacyl PEs from donor liposomes to acceptor liposomes, consisting of 1,2-dioleoyl-sn-glycero-3-phosphocholine, cholesterol and N-((6-biotinoyl)amino)hexanoyl)-1,2-distearoyl-sn-glycero-3-phosphoethanolamine (54.9:45:0.1 mole ratio), was measured. Donor liposomes were prepared using 1,2-distearoyl-sn-glycero-3-phosphocholine (50 mol%), cholesterol (45 mol%) and 5 mol% of either PEG-derivatized 1,2-dimyristoyl-sn-glycero-3-phosphoethanolamine (DMPE-PEG(2000)), 1,2-dipalmitoyl-sn-glycero-3-phosphoethanolamine (DPPE-PEG(2000)), or 1,2-distearoyl-sn-glycero-3-phosphoethanolamine (DSPE-PEG(2000)). Transfer of DSPE-PEG(2000) to the donor liposomes was not detected under the conditions employed. In contrast, DMPE-PEG(2000) was transferred efficiently even at 4 degrees C. Using an acceptor to donor liposome ratio of 1:4, the time required for DMPE-PEG(2000) to become evenly distributed between the two liposome populations (T(EQ)) at 4 degrees C and 37 degrees C was approx. 2 and <0.5 h, respectively. An increase in acyl chain length from C14:0 to C16:0 of the PEG-lipid resulted in a significant reduction in the rate of transfer as measured by this assay. The transfer of PEG-lipid out of biotinylated liposomes was also studied in mice following intravenous administration. The relative rates of transfer for the various PEG-lipids were found to be comparable under in vivo and in vitro conditions. These results suggest that it is possible to design targeted liposomes with the targeting ligand protected while in the circulation through the use of PEG-lipids that are selected on the basis of exchange characteristics which result in exposure of the shielded ligand following localization within a target tissue.

Animals↗
PubMed2001

Calmodulin-binding protein BP-10, a probable new member of plant nonspecific lipid transfer protein superfamily.

CaMBP-10 is a novel plant endogenous calmodulin-binding protein with important physiological functions. The partial cDNA sequence of this protein was cloned using RT-PCR. The deduced peptide (designated PCBP10) is composed of 74 amino acid residues containing a basic amphiphilic alpha-helix typical for calmodulin-binding proteins. PCBP10 shows very high amino acid sequence homology with plant nonspecific lipid-transfer proteins (nsLTPs). Sequence analysis also reveals that PCBP10 has similar amino acid composition to plant nsLTPs, and seven of the eight conserved cysteine residues are found in PCBP10. Furthermore, the secondary structure features of PCBP10 are very similar to those of plant nsLTPs. In addition, there are striking resemblances between CaMBP-10 and plant nsLTPs in their biochemical and physical properties. Our results suggest that CaMBP-10 is a novel member of the plant and nsLTP gene family, and the Ca(2+)/CaM regulative system may also play roles in lipid metabolism, defense reactions, and the adaptation of plants to natural environment.

Base Sequence↗
PubMed2001

Mitochondria are selectively eliminated from eukaryotic cells after blockade of caspases during apoptosis.

Pan caspase inhibitors are potentially powerful cell-protective agents that block apoptosis in response to a wide variety of insults that cause tissue degeneration. In many conditions, however, the blockade of apoptosis by caspase inhibitors does not permit long-term cell survival, but the reasons are not entirely clear. Here we show that the blockade of apoptosis by Boc.Aspartyl(O-methyl)CH2F can result in the highly selective elimination of the entire cohort of mitochondria, including mitochondrial DNA, from both neurons and HeLa cells, irrespective of the stimulus used to trigger apoptosis. In cells that lose their mitochondria, the nuclear DNA, Golgi apparatus, endoplasmic reticulum, centrioles, and plasma membrane remain undamaged. The capacity to remove mitochondria is both specific and regulated since mitochondrial loss in neurons is completely prevented by the expression of the antiapoptotic protein Bcl-2 and partially suppressed by the autolysosomal inhibitor bafilomycin. Cells without mitochondria are more tolerant to an anaerobic environment but are essentially irreversibly committed to death. Prevention of mitochondrial loss may be crucial for the long-term regeneration of tissues emerging from an apoptotic episode in which death was prevented by caspase blockade.

Amino Acid Chloromethyl Ketones↗
PubMed2001

Role of phosphatidylinositol 3-kinase-Akt pathway in nucleophosmin/anaplastic lymphoma kinase-mediated lymphomagenesis.

The NPM/ALK fusion gene, formed by the t(2;5) translocation in a subset of anaplastic large cell lymphomas, encodes a Mr 75,000 hybrid protein that contains the NH2-terminal portion of the nucleolar phosphoprotein nucleophosmin (NPM) joined to the entire cytoplasmic portion of the receptor tyrosine kinase anaplastic lymphoma kinase (ALK). NPM/ALK encodes a constitutively activated tyrosine kinase that belongs to the family of tyrosine kinases activated by chromosomal translocations. Our studies showed that NPM/ALK, similar to other members of this family, activates phosphatidylinositol 3-kinase (PI3K) and its downstream effector, serine/threonine kinase (Akt). PI3K was found in complex with NPM/ALK. Both PI3K and Akt kinase were permanently activated in NPM/ALK-transfected BaF3 murine hematopoietic cells and in NPM/ALK-positive, but not in NPM/ALK-negative, patient-derived anaplastic large cell lymphoma cell lines. In addition, Akt was phosphorylated/activated in protein samples isolated from four patients diagnosed with ALK-positive T/null-cell lymphomas. The PI3K inhibitors wortmannin and LY294002 induced apoptosis in NPM/ALK+ cells but exerted only minor effects on the control BaF3 parental cells and peripheral blood mononuclear cells stimulated by growth factors. Furthermore, retroviral infection of NPM/ALK+ BaF3 cells with a dominant-negative PI3K mutant (delta p85) or a dominant-negative Akt mutant (K179M) inhibited proliferation and clonogenic properties of the infected cells. Finally, the Akt mutant (K179M) suppressed the tumorigenicity of NPM/ALK-transfected BaF3 cells injected into syngeneic mice. In conclusion, our data indicate that NPM/ALK constitutively activates the PI3K-Akt pathway and that this pathway plays an important role in the NPM/ALK-mediated malignant transformation.

Animals↗
PubMed2001

Influence of P450 3A4 SRS-2 residues on cooperativity and/or regioselectivity of aflatoxin B(1) oxidation.

The major human liver drug-metabolizing cytochrome P450 enzymes P450 3A4 and P450 3A5 share >85% amino acid sequence identity yet exhibit different regioselectivity toward aflatoxin B(1) (AFB(1)) biotransformation [Gillam et al. (1995) Arch. Biochem. Biophys. 317, 74-384]. P450 3A4 prefers AFB1 3alpha-hydroxylation, which detoxifies and subsequently eliminates the hepatotoxin, over AFB1 exo-8,9-oxidation. P450 3A5, on the other hand, is a relatively sluggish 3alpha-hydroxylase and converts AFB(1) predominantly to the genotoxic exo-8,9-epoxide. Using a combination of approaches (sequence alignment, homology modeling and site-directed mutagenesis), we have previously identified several divergent residues in four of the six putative substrate recognition sites (SRSs) of P450 3A4, which when replaced individually with the corresponding amino acid of P450 3A5, resulted in a significant switch of the characteristic P450 3A4 AFB(1) regioselectivity toward that of P450 3A5 [Wang et al. (1998) Biochemistry 37, 12536-12545]. In particular, residues N206 and L210 in SRS-2 were found to be critical for AFB(1) detoxification via 3alpha-hydroxylation, and the corresponding mutants N206S and L210F most closely mimicked P450 3A5, not only in its regioselectivity of AFB(1) metabolism but also in its overall functional capacity. We have now further explored the plausible reasons for such relative inactivity of the SRS-2 mutants by examining N206S and additional mutants (L210A, L211F, L211A, and N206E) and found that the dramatically lowered activities of the N206S mutant are accompanied by a loss of cooperativity of AFB(1) oxidation. Molecular dynamics analyses with an existing P450 3A4 homology model [Szklarz and Halpert (1997) J. Comput. Aided Mol. Des. 11, 265] suggested that N206 (helix F) interacts with E244 (helix G), creating a salt bridge that stabilizes the protein structure and/or defines the active site cavity. To examine this possibility, several E244 mutants (E244A, V, N, S) were tested, of which E244S was the most notable for its relatively greater impairment of P450 3A4-dependent AFB(1) 3alpha-hydroxylation. However, the results with these E244 mutants failed to validate the N206-E244 interaction predicted from these molecular dynamics analyses. Collectively, our findings to date have led us to reconsider our original interpretations and to reexamine them in the light of AFB(1) molecular modeling analyses with a newly refined P450 3A4 homology model. These analyses predicted that F304 in SRS-4 (I-helix) plays a pivotal role in AFB(1) binding at the active site in either orientation leading to 3alpha- or exo-8,9-oxidation. Consistent with this prediction, conversion of F304 to Ala abolished P450 3A4-dependent AFB(1) 3alpha-hydroxylation and exo-8,9-oxidation.

Aflatoxin B1↗
PubMed2001

Construction and biological characterization of an HB-GAM/FGF-1 chimera for vascular tissue engineering.

OBJECTIVE: Cardiovascular tissue engineering approaches to vessel wall restoration have focused on the potent but relatively nonspecific and heparin-dependent mesenchymal cell mitogen fibroblast growth factor 1 (FGF-1). We hypothesized that linking FGF-1 to a sequence likely to bind to cell surface receptors relatively more abundant on endothelial cells (ECs) might induce a relative greater EC bioavailability of the FGF-1. We constructed a heparin-binding growth-associated molecule (HB-GAM)/FGF-1 chimera by linking full-length human HB-GAM to the amino-terminus of human FGF-1beta (21-154) and tested its activities on smooth muscle cells (SMCs) and ECs. METHODS: Primary canine carotid SMCs and jugular vein ECs were plated in 96-well plates in media containing 10% fetal bovine serum and grown to approximately 80% confluence. After being growth arrested in serum-free media for 24 hours, the cells were exposed to concentration ranges of cytokines and heparin, and proliferation was measured with tritiated-thymidine incorporation. Twenty percent fetal bovine serum was used as positive control, and phosphate-buffered saline was used as negative control. RESULTS: In the presence of heparin the HB-GAM/FGF-1 chimera stimulated less SMC proliferation than did the wild-type FGF-1 with a median effective dose of approximately 0.3 nmol versus approximately 0.1 nmol (P <.001). By contrast, the chimera retained full stimulating activity on EC proliferation with a median effective dose of 0.06 nmol for both cytokines. Unlike the wild-type protein, the chimera possessed heparin-independent activity. In the absence of heparin, the chimera induced dose-dependent EC and SMC proliferation at 0.06 nmol or more compared with the wild-type FGF-1, which stimulated minimal DNA synthesis at 6.0-nmol concentrations. CONCLUSIONS: The HB-GAM/FGF-1 chimera displays significantly greater and uniquely heparin-independent mitogenic activity for both cell types, and in the presence of heparin it displays a significantly greater EC specificity.

Animals↗
PubMed2001

Multiple protein functions of paired in Drosophila development and their conservation in the Gooseberry and Pax3 homologs.

The Drosophila segmentation gene paired, whose product is homologous to the Drosophila Gooseberry and mammalian Pax3 proteins, has three general functions: proper development of the larval cuticle, survival to adulthood and male fertility. Both DNA-binding domains, the conserved N-terminal paired-domain and prd-type homeodomain, are required within the same molecule for all general paired functions, whereas a conserved His-Pro repeat located near its C terminus is a transactivation domain potentiating these functions. The C-terminal moiety of Paired includes two additional functional motifs: one, also present in Gooseberry and Pax3, is required for segmentation and cuticle development; the other, retained only in Gooseberry, is necessary for survival. The male fertility function, which cannot be replaced by Gooseberry and Pax3, is specified by the conserved N-terminal rather than the divergent C-terminal moiety of Paired. We conclude that the functional diversification of paired, gooseberry and Pax3, primarily determined by variations in their enhancers, is modified by adaptations of their coding regions as a necessary consequence of their newly acquired spatiotemporal expression.

Amino Acid Motifs↗
PubMed2000

Inhibition of JNK by overexpression of the JNL binding domain of JIP-1 prevents apoptosis in sympathetic neurons.

Studies in non-neuronal cells show that c-Jun N-terminal kinases (JNK) play a key role in apoptotic cell death. In some neurons JNK is also thought to initiate cell death by the activation of c-Jun. JNK inhibition has been achieved pharmacologically by inhibiting upstream kinases, but there has been no direct demonstration that inhibition of JNK can prevent neuronal death. We have therefore examined whether the JNK binding domain (JBD) of JNK-interacting protein-1 (JIP-1, a scaffold protein and specific inhibitor of JNK) can inhibit c-Jun phosphorylation and support the survival of sympathetic neurons deprived of NGF. We show that expression of the JBD in >80% of neurons was sufficient to prevent the phosphorylation of c-Jun and its nuclear accumulation as well as abrogate neuronal cell death induced by NGF deprivation. JBD expression also preserved the capacity of mitochondria to reduce MTT. Interestingly, although the PTB domain of JIP was reported to interact with rhoGEF, expression of the JBD domain was sufficient to localize the protein to the membrane cortex and growth cones. Hence, JNK activation is a key event in apoptotic death induced by NGF withdrawal, where its point of action lies upstream of mitochondrial dysfunction.

Adaptor Proteins, Signal Transducing↗
PubMed2000

Death commitment point is advanced by axotomy in sympathetic neurons.

Axotomized neurons have several characteristics that are different from intact neurons. Here we show that, unlike established cultures, the axotomized sympathetic neurons deprived of NGF become committed to die before caspase activation, since the same proportion of NGF-deprived neurons are rescued by NGF regardless of whether caspases are inhibited by the pan-caspase inhibitor Boc-Asp(O-methyl)-CH(2)F (BAF). Despite prolonged Akt and ERK signaling induced by NGF after BAF treatment has prevented death, the neurons fail to increase protein synthesis, recover ATP levels, or grow. Within 3 d, all the mitochondria disappear without apparent removal of any other organelles or loss of membrane integrity. Although NGF does rescue intact BAF-treated 6-d cultures after NGF deprivation, rescue by NGF fails when these neurons are axotomized before NGF deprivation and BAF treatment. Moreover, cytosolic cytochrome c rapidly kills axotomized neurons. We propose that axotomy induces signals that make sympathetic neurons competent to die prematurely. NGF cannot repair these NGF-deprived, BAF-treated neurons because receptor signaling (which is normal) is uncoupled from protein renewal, and the mitochondria (which are damaged) go on to be eliminated. Hence, the order of steps underlying neuronal death commitment is mutable and open to regulation.

Adenosine Triphosphate↗
PubMed2000

Drosophila female sexual behavior induced by sterile males showing copulation complementation.

Females of most animal species are usually inseminated by more than one male, which allows sperm from different males to compete for fertilization. To prevent invasion of sperm from other males, Drosophila males elicit a rejection behavior in their mates after copulation. Using paired mutant males that, for the lack of accessory glands, are sterile, we show that this rejection behavior is induced exclusively by the secreted accessory gland products transferred to the female during copulation. Moreover, the activities of sperm and accessory gland products are complementary and interdependent: both sperm fertility and rejection behavior depend on accessory gland products whose prolonged activities, in turn, require the presence of sperm. Fertility of sperm from paired males can be restored by accessory gland products of spermless males in "copulation complementation" experiments. Our observations may have important implications for the role of sexual behavior in evolution and for the treatment of male sexual dysfunction in humans.

Animals↗
PubMed2000

The Ras/phosphatidylinositol 3-kinase and Ras/ERK pathways function as independent survival modules each of which inhibits a distinct apoptotic signaling pathway in sympathetic neurons.

Ras promotes robust survival of many cell systems by activating the phosphatidylinositol 3-kinase (PI3-kinase)/Akt pathway, but little is understood about the survival functions of the Ras/ERK pathway. We have used three different effector-loop mutant forms of Ras, each of which activates a single downstream effector pathway, to dissect their individual contributions to survival of nerve growth factor (NGF)-dependent sympathetic neurons. The PI3-kinase pathway-selective protein Ras(Val-12)Y40C was as powerful as oncogenic Ras(Val-12) in preventing apoptosis induced by NGF deprivation but conferred no protection against apoptosis induced by cytosine arabinoside. Identical results were obtained with transfected Akt. In contrast, the ERK pathway-selective protein Ras(Val-12)T35S had no protective effects on NGF-deprived neurons but was almost as strongly protective as Ras(Val-12) against cytosine arabinoside-induced apoptosis. The protective effects of Ras(Val-12)T35S against cytosine arabinoside were completely abolished by the ERK pathway inhibitor PD98059. Ras(Val-12)E37G, an activator of RalGDS, had no survival effect on either death pathway, similar to RasS17N, the full survival antagonist. Thus, Ras provides two independent survival pathways each of which inhibits a distinct apoptotic mechanism. Our study presents one of the few clear-cut cases where only the Ras/ERK, but not the Ras/PI3K/Akt pathway, plays a dominant survival signaling role.

Animals↗
PubMed2000

Carbon monoxide and nitric oxide as coneurotransmitters in the enteric nervous system: evidence from genomic deletion of biosynthetic enzymes.

Nitric oxide (NO) and carbon monoxide (CO) seem to be neurotransmitters in the brain. The colocalization of their respective biosynthetic enzymes, neuronal NO synthase (nNOS) and heme oxygenase-2 (HO2), in enteric neurons and altered intestinal function in mice with genomic deletion of the enzymes (nNOS(Delta/Delta) and HO2(Delta/Delta)) suggest neurotransmitter roles for NO and CO in the enteric nervous system. We now establish that NO and CO are both neurotransmitters that interact as cotransmitters. Small intestinal smooth muscle cells from nNOS(Delta/Delta) and HO2(Delta/Delta) mice are depolarized, with apparent additive effects in the double knockouts (HO2(Delta/Delta)/nNOS(Delta/Delta)). Muscle relaxation and inhibitory neurotransmission are reduced in the mutant mice. In HO2(Delta/Delta) preparations, responses to electrical field stimulation are nearly abolished despite persistent nNOS expression, whereas exogenous CO restores normal responses, indicating that the NO system does not function in the absence of CO generation.

Animals↗
PubMed2000

The cysteine-free fibroblast growth factor 1 mutant induces heparin-independent proliferation of endothelial cells and smooth muscle cells.

BACKGROUND: The structure/function relationships of fibroblast growth factor 1 (FGF-1) are being investigated using site mutation, yielding novel structures with potential clinical applicability for modulating tissue responses to vascular interventions. We generated a mutant FGF-1 in which all three cysteines were converted to serines and then tested the relative mitogenic activities on endothelial cells (ECs) and smooth muscle cells (SMCs) and the molecular stability of the protein to thrombin-induced degradation. METHODS: The dose responses of wild-type FGF-1 and the Cys-free mutant in the absence or presence of heparin were tested on ECs and SMCs. Cell proliferation was measured by [(3)H]thymidine incorporation. Data were normalized by positive control (20% fetal bovine serum) and expressed as percentage of positive control for comparison. The molecular stability was examined by exposure of the cytokines to thrombin at 37 degrees C for 0.5-24 h and then analyzed by sodium dodecyl sulfate-polyacrylamide gel electrophoresis. RESULTS: Unlike wild-type FGF-1 which induced only minimal DNA synthesis at concentrations as high as 100 ng/ml, the Cys-free mutant induced a dose-dependent proliferation starting at 1 ng/ml on both ECs and SMCs in the absence of heparin. At 100 ng/ml, Cys-free mutant induced 4-fold more proliferation than wild-type FGF-1 on ECs (76.64 +/- 13.39% vs 14.58 +/- 1.38%, P < 0.01) and 12-fold more proliferation on SMCs (143.52 +/- 9.96 vs 11.25 +/- 3.32, P < 0.01). Heparin 5 U/ml potentiated the mitogenic activity of the Cys-free mutant at low dose range. Both proteins were degraded by thrombin progressively. But the Cys-free mutant showed more susceptibility with accelerated appearance of lower-molecular-weight fragment bands after incubation with thrombin. CONCLUSIONS: Conversion of cysteine residues to serine changed the heparin dependency of the growth factor and increased its mitogenic activity and its susceptibility to thrombin-induced degradation.

Animals↗
PubMed2000

Evolution of Hoxa-11 in lineages phylogenetically positioned along the fin-limb transition.

HOXA11 is a transcription factor implicated in paired appendage development. To identify signatures of evolutionary change in the structural, and putative functional, domains of HOXA11, we studied its evolution in tetrapod and nontetrapod lineages that represent approximately 1.5 billion years of evolutionary time. Here, Hoxa-11 gene proper sequences were determined for frog (Xenopus tropicalis), coelacanth (Latimeria chalumnae), common zebrafish (Danio rerio; Hoxa-11a and Hoxa-11b paralogs), and giant zebrafish (D. aequipinnatus; Hoxa-11b) and aligned against previously published Hoxa-11 sequences of human, mouse, chick, and newt. Based on aligned Hoxa-11 amino acid sequences, the protein was demarcated into three segments: Domains I (N-terminal) and III (homeobox + C-terminal), which varied slightly in rates and patterns of evolution, and a variable, overall hydrophilic region (HyD), which partially overlaps with Domain I. As judged by character reconstructions of HOXA11 Domains I and III, no significant changes in rates of coding sequence evolution occurred in tetrapods (frog and chick), relative to coelacanth (a lobe-finned fish), i.e., across the fin-limb transition. Accelerated rates of Hoxa-11 coding sequence evolution were observed for the mammalian and newt lineages. This was shown to be a gene-specific phenomenon. The duplicated Hoxa-11a and Hoxa-11b genes of zebrafish exhibited accelerated rates of evolution and accumulated substitutions at sites that are conserved among coelacanth and all tetrapods examined. Amino acid sequence comparisons of the HyD of HOXA11 suggested that a putative repressor subdomain, containing stretches of consecutive alanine residues, emerged within the tetrapods. A high degree of nucleotide conservation in the 5' half of the Hoxa-11 intron was observed for tetrapod and nontetrapod lineages. Using electrophoretic mobility shift assays, a 35-bp intron sequence, which is 100% conserved in all Hoxa-11 loci except for the zebrafish Hoxa-11a paralog, was found to bind protein(s) in HeLa and chick whole-cell extracts.

Alanine↗
PubMed2000

The S130K fibroblast growth factor-1 mutant induces heparin-independent proliferation and is resistant to thrombin degradation in fibrin glue.

OBJECTIVE: Site-directed mutagenesis is an important technique that can alter cytokine function, thereby eliciting desired responses. S130K is a mutation of fibroblast growth factor-1 (FGF-1), with lysine replacing serine in the heparin-binding site. We measured molecular stability and mitogenic activity of FGF-1 and S130K, both in the media and when suspended in fibrin glue (FG), on smooth muscle cells (SMCs) and endothelial cells (ECs) to determine if the mutation altered the function and potential clinical applicability. METHODS: EC and SMC proliferation of soluble FGF-1 or S130K at 0, 0. 1, 1, 10, or 100 ng/mL with heparin at 0, 5, 50, or 500 units (U)/mL was measured on growth-arrested cells in serum-free media. EC and SMC proliferation assays with cells on FG containing either FGF-1 or S130K at 0, 1, 10, 100, or 1000 ng/mL in combination with heparin at 0, 5, 50 or 500 U/mL were also performed during the exponential growth phase. Molecular degradation by thrombin was measured by sodium dodecylsulfate-polyacrylamide gel electrophoresis. RESULTS: S130K induces greater EC and SMC proliferation in the absence of heparin than FGF-1 does (P <.0001 for both the 10 and 100 ng/mL doses). S130K is also significantly more potent than FGF-1 in the presence of heparin. Heparin in the media enhances cytokine-induced SMC and EC proliferation at doses of 5 U/mL, but inhibits SMC proliferation at concentrations of 500 U/mL. For the FG data, unlike FGF-1, S130K induces EC and SMC proliferation in the absence of heparin. The addition of 5 U/mL of heparin enhances the proliferation induced by S130K. For ECs, as the heparin dose increases to 50 U/mL, proliferation decreases, as compared with the 5 U/mL concentration when either FGF-1 or S130K in the FG was compared at concentrations of 10, 100, and 1000 ng/mL (P <.01). S130K is more potent in FG than is FGF-1 both with and without heparin and exhibits maximal EC and SMC proliferation at 10 ng/mL, whereas FGF-1 activity is maximal at 100 ng/mL. Gel electrophoresis demonstrated that S130K was relatively more resistant to thrombin degradation than FGF-1. CONCLUSIONS: Site-directed mutagenesis changed the potency and the heparin dependency on cellular proliferation of FGF-1 in vitro. These techniques should allow the delivery of mutant growth factors to areas of vascular intervention to induce specific, desired responses. We believe that these studies will enhance our knowledge of the function of various regions of the FGF-1 molecule, allowing us to more precisely design increasingly more useful FGF-1 mutants.

Animals↗
PubMed2000

Combinatorial preferences affect molecular similarity/diversity calculations using binary fingerprints and Tanimoto coefficients

A combinatorial method was developed to calculate complete distributions of the Tanimoto coefficient (Tc) for binary fingerprint (FP) representations of specified length, regardless of the chemical parameters they reflect. Theoretical Tc distributions were calculated for FPs consisting of up to 67 bit positions which revealed significant statistical preferences of certain Tc values. Calculation of Tc distributions in a large compound database using different FPs mirrored the effects identified by our general analysis. On the basis of these findings, an average Tc is biased by statistically preferred values.

Journal Article↗
PubMed2000

Searching for molecules with similar biological activity: analysis by fingerprint profiling.

We have recently developed a mini-fingerprint (MFP) representation for small molecules that performs well in database searches for compounds with similar biological activity. The MFP consists of only 54 bit positions that account for numerical ranges of three two-dimensional (2D) descriptors or the presence or absence of defined structural fragments. Here we present an analysis method, termed fingerprint profiling, to systematically compare bit patterns of compounds belonging to different biological activity classes. Some but not all bit positions were variably occupied in seven different activity classes and responsible for the detection of structure-activity differences. The analysis has made it possible to rank bit positions and encoded molecular descriptors according to their importance for our similarity search calculations. Fingerprint profiling can be applied to any keyed bit string representation and should be helpful, for example, to analyze descriptor distributions in large compound databases.

Computer-Aided Design↗