Dimethyl Suberimidate Research Articles

Pro-prion Binds Filamin A, Facilitating Its Interaction with Integrin β1, and Contributes to Melanomagenesis

Filamin A (FLNA) is an integrator of cell mechanics and signaling. The spreading and migration observed in FLNA sufficient A7 melanoma cells but not in the parental FLNA deficient M2 cells have been attributed to FLNA. In A7 and M2 cells, the normal prion (PrP) exists as pro-PrP, retaining its glycosylphosphatidyl-inositol (GPI) anchor peptide signal sequence (GPI-PSS). The GPI-PSS of PrP has a FLNA binding motif and binds FLNA. Reducing PrP expression in A7 cells alters the spatial distribution of FLNA and organization of actin and diminishes cell spreading and migration. Integrin β1 also binds FLNA. In A7 cells, FLNA, PrP, and integrin β1 exist as two independent, yet functionally linked, complexes; they are FLNA with PrP or FLNA with integrin β1. Reducing PrP expression in A7 cells decreases the amount of integrin β1 bound to FLNA. A PrP GPI-PSS synthetic peptide that crosses the cell membrane inhibits A7 cell spreading and migration. Thus, in A7 cells FLNA does not act alone; the binding of pro-PrP enhances association between FLNA and integrin β1, which then promotes cell spreading and migration. Pro-PrP is detected in melanoma in situ but not in melanocyte. Invasive melanoma has more pro-PrP. The binding of pro-PrP to FLNA, therefore, contributes to melanomagenesis.

Cross-linking with bifunctional reagents and its application to the study of the molecular symmetry and the arrangement of subunits in hexameric protein oligomers

Cross-linking with a bifunctional reagent and subsequent SDS gel electrophoresis is a simple but effective method to study the symmetry and arrangement of subunits in oligomeric proteins. In this study, theoretical expressions for the description of cross-linking patterns were derived for protein homohexamers through extension of the method used for tetramers by Hajdu et al. (1976). The derived equations were used for the analysis of cross-linking by glutardialdehyde of four protein hexamers: beef liver glutamate dehydrogenase (GDH), jack bean urease, hemocyanin from the spiny lobster Panulirus pencillatus ( PpHc), Escherichia coli glutamate decarboxylase (GDC) and for analysis of published data on the cross-linking of hexameric E. coli rho by dimethyl suberimidate. Best fit models showed that the subunits in the first four proteins are arranged according to D 3 symmetry in two layers, each subunit able to cross-link to three neighboring subunits for GDH and urease, or to four for PpHc and GDC. The findings indicate a dimer-of-trimers eclipsed arrangement of subunits for GDH and urease and a trimer-of-dimers staggered one for PpHc and GDC. In rho, the subunits are arranged according to D 3 symmetry in a trimer-of-dimers ring. The conclusions from cross-linking of GDH and GDC, PpHc and rho are consistent with results from X-ray crystal structure, those for urease with findings from electron microscopy.

Structural Basis for the Recognition of Histone H4 by the Histone-Chaperone RbAp46

SummaryRbAp46 and RbAp48 (pRB-associated proteins p46 and p48, also known as RBBP7 and RBBP4, respectively) are highly homologous histone chaperones that play key roles in establishing and maintaining chromatin structure. We report here the crystal structure of human RbAp46 bound to histone H4. RbAp46 folds into a seven-bladed β propeller structure and binds histone H4 in a groove formed between an N-terminal α helix and an extended loop inserted into blade six. Surprisingly, histone H4 adopts a different conformation when interacting with RbAp46 than it does in either the nucleosome or in the complex with ASF1, another histone chaperone. Our structural and biochemical results suggest that when a histone H3/H4 dimer (or tetramer) binds to RbAp46 or RbAp48, helix 1 of histone H4 unfolds to interact with the histone chaperone. We discuss the implications of our findings for the assembly and function of RbAp46 and RbAp48 complexes.

An AFM study on mechanical properties of native and dimethyl suberimidate cross‐linked pericardium tissue

Changes in the stiffness of hog pericardium tissue, native and treated with dimethyl suberimidate (DMS), are investigated by atomic force microscopy (AFM). Young's modulus is calculated on the basis of the Hertz-Sneddon model. The cross-linking process increases the stiffness of the tissue. The values of Young's modulus are higher for the DMS stabilized pericardium than for the native one. We also observe that the Young's modulus of native tissue increases when the time between getting the biological material and performing the measurements is longer. This process is probably connected with natural degradation of the biological samples.

Cytotoxicity of Polyspermine-Ribonuclease A and Polyspermine-Dimeric Ribonuclease A

Polyspermine-ribonuclease A (PS-RNase A) and polyspermine-dimeric ribonuclease A (PS-dimeric RNase A) were prepared by cross-linking ribonuclease A or its covalently linked dimer to polyspermine (PS) using dimethyl suberimidate. The two RNase A derivatives were tested for a possible antitumor action. The in vitro and in vivo cytotoxic activity of PS-RNase A, although strong, is not higher than that known for free polyspermine. PS-dimeric RNase A, which was characterized by mass spectroscopy, titration of free amine groups, and enzymatic assays, proved instead to be a definitely more efficient antitumor agent, both in vitro and in vivo. This result could tentatively be explained in view of the importance of positive charges for ribonuclease activity, considering the higher basicity of PS-dimeric RNase A compared to that of PS-(monomeric)RNase A. It must be also taken into account that the dimeric RNase A moiety of PS-dimeric RNase A could evade the cytoplasmic ribonuclease inhibitor, which instead could trap the monomeric RNase A moiety of the other derivative. The two RNase A derivatives degrade poly(A).poly(U) under conditions where native RNase A is inactive. The results of this work demonstrate once again the importance of positive charges for the functions of mammalian pancreatic type ribonucleases in general, in particular for RNase A derivatives, and the potential therapeutic use of the ribonuclease A derivatives.

Ss-LrpB from Sulfolobus solfataricus Condenses about 100 Base Pairs of Its Own Operator DNA into Globular Nucleoprotein Complexes

Ss-LrpB from the hyperthermoacidophilic crenarchaeote Sulfolobus solfataricus P2 is a member of the Lrp-like family of Bacterial/Archaeal transcription regulators that binds its own control region at three regularly spaced and partially conserved 15-bp-long imperfect palindromes. We have used atomic force microscopy to analyze the architecture of Ss-LrpB.DNA complexes with a different stoichiometry formed with the wild type operator and with an operator mutant. Binding of dimeric Ss-LrpB to all three target sites is accompanied by the formation of globular complexes, in which the protein induces strong DNA deformations. Furthermore, DNA contour length foreshortening of these complexes indicates DNA wrapping, with about 100 bp being condensed. The average bending angle is 260 degrees . The establishment of protein-protein contacts between Ss-LrpB dimers in these globular complexes will contribute to the cooperativity of the binding. The profound remodeling of the control region is expected to have a strong impact on gene expression and might constitute the key element in the autoregulatory process.

Mutational, structural and functional studies of the mitochondrial translational initiation factor 2 on the mammalian mitochondrial ribosome

The mammalian mitochondrial ribosome translates only 13 gene products from the mitochondrial genome. Initiation of translation of these proteins requires mitochondrial translational initiation factor 2 (IF-2mt), a G-protein. Mutational analysis of the X residue (Cys residue 290) in the NKXD guanine ring binding motif in the GTPase center of IF-2mt yielded significant functional changes. The mutation cysteine to serine (C290S) had very little effect on the function of this protein but the C290A and C290I mutations both resulted in a protein that was unable to fold and subsequently degraded. Interestingly, although the X residue of this motif is not thought to participate in the specific binding and GTPase activity of IF-2mt, mutation to even the most highly conserved residue in IF-2 at the X position, Ile, was detrimental to the protein’s structure and function. IF-2mt is known to specifically bind to the 28 S small subunit of the 55 S mammalian mitochondrial ribosome with nanomolar affinity. CryoEM studies on the 55 S mitochondrial ribosome indicate that the small subunit has a number of interesting features. Proteomic studies have shown that there are 29 proteins in the small subunit only half of which have homologs in bacteria. The binding site of IF-2mt on the 28 S subunit is being examined using dimethyl suberimidate (DMS) to crosslink ribosomal proteins to IF-2mt. The crosslinked proteins will be identified using mass spectrometry. These studies will yield important clues to the structure and mechanism of action of the mammalian mitochondrial ribosome. Funding: NIH GM32734.

Nuclear Transport of Trimeric Assembly Intermediates Exerts a Morphogenetic Control on the Icosahedral Parvovirus Capsid

The connection between nuclear transport and morphogenesis of a large macromolecular entity has been investigated using the karyophylic capsid of the parvovirus minute virus of mice (MVM) as a model. The VP1 (82 kDa) and VP2 (63 kDa) proteins forming the T = 1 icosahedral MVM capsid at the respective 1:5 molar ratio of synthesis, could be covalently cross-linked with dimethyl suberimidate into two types of oligomeric assemblies, which were present at stoichiometric amounts in infected cell extracts and purified viral particles. The larger species contained VP1 and corresponded in size (200 kDa) to a heterotrimer of one VP1 and two VP2 subunits. The smaller species contained VP2 only and corresponded in size (180 kDa) to a homotrimer. The introduction of bulky residues or the truncation of side-chains involved in multiple interactions at the interfaces between trimers of VPs in the MVM capsid, produced the accumulation of trimeric intermediates that were competent in nuclear translocation but not in capsid assembly. These results indicate that MVM maturation proceeds by cytoplasmic oligomerization of the capsid subunits into two types of trimers, which are the assembly intermediates competent to translocate across the nuclear membrane. Consistent with this conclusion, mutations at basic residues that inactivate a previously identified beta-stranded nuclear localization motif, which notably are not involved in inter or intra-subunit contacts, led to cytoplasmic retention of the two types of trimers, with no evidence for other assembly intermediates. Although a fraction of the VP1-containing trimers were translocated into the nucleus driven by the conventional nuclear transport signal of VP1 N terminus, their further assembly in the absence of the VP2-only trimers yielded large molecular mass amorphous aggregates. Therefore, the nuclear transport stoichiometry of assembly intermediates may exert a morphogenetic quality control on macromolecular complexes like the MVM capsid.

Atomic force microscopy investigation of chemically stabilized pericardium tissue

Native and chemically stabilized porcine pericardium tissue was imaged by the contact mode atomic force microscopy (AFM), in air. Chemically stabilized pericardium is used as a tissue-derived biomaterial in various fields of the reconstructive and replacement surgery. Collagen type I is the main component of the fibrous layer of the pericardium tissue. In this study, the surface topography of collagen fibrils in their native state in tissue and after chemical stabilization with different cross-linking reagents: glutaraldehyde (GA), dimethyl suberimidate (DMS) and tannic acid (TA) was investigated. It has been found that chemical stabilization causes considerable changes in the surface topography of collagen fibrils as well as in the spatial organization of the fibrils within the tissue. The observed changes in the D-spacing pattern of the collagen fibril correspond to the formation of intrafibrilar cross-links, whereas formation of interfibrilar cross-links is mainly responsible for the observed tangled spatial arrangement of fibrils and crimp structure of the tissue surface. The crimp structure was distinctly seen for the GA cross-linked tissue. Surface heterogeneity of the cross-linking process was observed for the DMS-stabilized tissue. SDS-PAGE electrophoresis was performed in order to evaluate the stabilization effect of the tissues treated with the cross-linking reagents. It has been found that stabilization with DMS, GA or TA enhances significantly the tissue resistance to SDS/NaCl extraction. The relation between the tissue stability and changes in the topography of the tissue surface was interpreted in terms of different nature of cross-links formed by DMS, GA and TA with collagen.

Dimethyl suberimidate cross-linked pericardium tissue: Raman spectroscopic and atomic force microscopy investigations

Chemically stabilized pericardium tissue is widely used as a tissue-derived biomaterial for the preparation of bioprostheses such as heart valves or vascular grafts. The bifunctional imidoester dimethyl suberimidate (DMS) belongs to the wide class of the cross-linking reagents and is often used to cross-link a variety of proteins, including collagen matrices and collagen-based tissues. Raman spectroscopy in the wide frequency range 200–4000 cm −1 and contact mode atomic force microscopy (AFM) have been employed to investigate the structural changes and chemical bonds in DMS cross-linked porcine pericardium tissue. It has been found, that in addition to the commonly accepted reaction with the ε-amine groups of lysine or hydroxylysine residues, DMS may interact also with the carbonyl CO and amide NH groups of the peptide bond in collagen. Our paper presents for the first time spectral evidence for the peptide contribution to the formation of DMS–collagen cross-links. The results confirm also possible competition between the hydrolysis of the free imidoester group and cross-linking reaction. Products of the partial alkaline hydrolysis of DMS have been found in the spectra. The observed changes in the surface topography of the fibrils as well as in their spatial organization in the tissue support the formation of both intra- and interfibrillar cross-links in DMS-stabilized tissue.

A Novel Enzyme Complex of Orotate Phosphoribosyltransferase and Orotidine 5‘-Monophosphate Decarboxylase in Human Malaria Parasite Plasmodium falciparum: Physical Association, Kinetics, and Inhibition Characterization,

Human malaria parasite, Plasmodium falciparum, can only synthesize pyrimidine nucleotides using the de novo pathway, whereas mammalian cells obtain pyrimidine nucleotides from both the de novo and salvage pathways. The parasite's orotate phosphoribosyltransferase (PfOPRT) and orotidine 5'-monophosphate decarboxylase (PfOMPDC) of the de novo pyrimidine pathway are attractive targets for antimalarial drug development. Previously, we have reported that the two enzymes in P. falciparum exist as a multienzyme complex containing two subunits each of 33-kDa PfOPRT and 38-kDa PfOMPDC. In this report, the gene encoding PfOPRT has been cloned and expressed in Escherichia coli. An open reading frame of PfOMPDC gene was identified in the malaria genome database, and PfOMPDC was cloned from P. falciparum cDNA, functionally expressed in E. coli, purified, and characterized. The protein sequence has <20% identity with human OMPDC and four microbial OMPDC for which crystal structures are known. Recombinant PfOMPDC was catalytically active in a dimeric form. Both recombinant PfOPRT and PfOMPDC monofunctional enzymes were kinetically different from the native multienzyme complex purified from P. falciparum. Oligomerization of PfOPRT and PfOMPDC cross-linked by dimethyl suberimidate indicated that they were tightly associated as the heterotetrameric 140-kDa complex, (PfOPRT)2(PfOMPDC)2. Kinetic analysis of the PfOPRT-PfOMPDC associated complex was similar to that of the native P. falciparum enzymes and was different from that of the bifunctional human enzymes. Interestingly, a nanomolar inhibitor of the yeast OMPDC, 6-thiocarboxamido-uridine 5'-monophosphate, was about 5 orders of magnitude less effective on the PfOMPDC than on the yeast enzyme. Our results support that the malaria parasite has unique structural and functional properties, sharing characteristics of the monofunctional pyrimidine-metabolizing enzymes in prokaryotes and bifunctional complexes in eukaryotes.

Highly Sensitive Immunoprecipitation Method for Extracting and Concentrating Low-Abundance Proteins from Human Serum

Purification and identification of small, low-abundance proteins from complex samples such as serum for use in Western blotting experiments has long been a challenge. Unfortunately, commercially available methods are not always suitable for small sample sizes and low-abundance proteins such as cardiac troponin T (cTnT). In this report we present a highly sensitive immunoprecipitation method we have developed for extracting and concentrating low-abundance proteins from serum. We compared the performance of our method with that of the commercially available ImmunoPure® Protein-A IgG orientation Kit (Pierce), using antibodies directed against cTnT and serum of a patient with acute myocardial infarction (AMI). Both methods are based on the binding of antibodies to Sepharose beads and subsequent cross-linking with a diimido ester. The formation of amidines between imido esters and amines from protein side chains was first described in 1962 by Hunter et al. (1), who suggested this for cross-linking purposes. Levy et al. (2) described the use of diimido esters for covalent coupling of antibodies to immobilized antigens. Later studies reported similar methods for antibody immobilization with cross-linkers such as dimethyl pimelimidate and dimethyl suberimidate (3)(4). These methods are based on the fact that an amine is able to react with an imido ester only if it is deprotonated, which occurs at pH > p K a. At these pH values, the nucleophilic attack of an R-NH2 on one of the two outer backbone carbon atoms of dimethyl pimelimidate produces a tetrahedral intermediate that splits into the amidine link and methanol. Hunter et al. (1) showed that the right choice of pH even allows a distinction to be made between α- and e-amines. They showed that a pH between 7 and 8 favors reaction with α-amines, whereas a pH between 9 and 10 favors reaction with e-amines. …

Human malaria parasite orotate phosphoribosyltransferase: functional expression, characterization of kinetic reaction mechanism and inhibition profile.

Plasmodium falciparum, the causative agent of the most lethal form of human malaria, relies on de novo pyrimidine biosynthesis. A gene encoding orotate phosphoribosyltransferase (OPRT), the fifth enzyme of the de novo pathway catalyzing formation of orotidine 5'-monophosphate (OMP) and pyrophosphate (PP(i)) from 5-phosphoribosyl-1-pyrophosphate (PRPP) and orotate, was identified from P. falciparum (pfOPRT). The deduced amino acid sequence for pfOPRT was compared with OPRTs from other organisms and found to be most similar to that of Escherichia coli. The catalytic residues and consensus sequences for substrate binding in the enzyme were conserved among other organisms. The pfOPRT was exceptional in that it contained a unique insertion of 20 amino acids and an amino-terminal extension of 66 amino acids, making the longest amino acid sequence (281 amino acids with a predicted molecular mass of 33kDa). The cDNA of the pfOPRT gene was cloned, sequenced and functionally expressed in soluble form. The recombinant pfOPRT was purified from the E. coli lysate by two steps, nickel metal-affinity and gel-filtration chromatography. From 1l E. coli culture, 1.2-1.5mg of pure pfOPRT was obtained. SDS-PAGE revealed that the pfOPRT had a molecular mass of 33kDa and analytical gel-filtration chromatography showed that the enzyme activity eluted at approximately 67kDa. Using dimethyl suberimidate to cross-link neighboring subunits of the pfOPRT, it was confirmed that the native enzyme exists in a dimeric form. The steady state kinetics of initial velocity and product inhibition studies indicate that the enzyme pfOPRT follows a random sequential kinetic mechanism. Compounds aimed at the pfOPRT nexus may act against the parasite through at least two mechanisms: by directly inhibiting the enzyme activity, or be processed to an inhibitor of thymidylate synthase. This study provides a working system with which to investigate new antimalarial agents targeted against P. falciparum OPRT.

Structural effects of carbohydrate-containing polycations on gene delivery. 3. Cyclodextrin type and functionalization.

Linear cationic beta-cyclodextrin (beta-CD)-based polymers can form polyplexes with plasmid DNA and transfect cultured cells. The effectiveness of the gene delivery and the cellular toxicity has been related to structural features in these polycations. Previous beta-CD polycations were prepared from the cocondensation of 6(A),6(D)-dideoxy-6(A),6(D)-diamino-beta-CD monomers with other difunctionalized monomers such as dimethyl suberimidate (DMS). Here, the type of CD and its functionalization are varied by synthesizing numerous 3(A),3(B)-dideoxy-3(A),3(B)-diamino-beta- and gamma-CD monomers. Both alkyl- and alkoxydiamines are prepared in order to vary the nature of the spacing between the CD and the primary amines in the monomers. These diamino-CD-monomers are polymerized with DMS to yield amidine-based polycations. The nature of the spacer between the CD-ring and the primary amines of each monomer is found to influence both molecular weight and polydispersity of the polycations. When these polycations are used to form polyplexes with plasmid DNA, longer alkyl regions between the CD and the charge centers in the polycation backbone increase transfection efficiency and toxicity in BHK-21 cells, while increasing hydrophilicity of the spacer (alkoxy versus alkyl) provides for lower toxicity. Further, gamma-CD-based polycations are shown to be less toxic than otherwise identical beta-CD-based polycations.

High-Efficiency Solid-Phase Capture Using Glass Beads Bonded to Microcentrifuge Tubes: Immunoprecipitation of Proteins from Cell Extracts and Assessment of Ras Activation

We have bonded glass microbeads (425–600 μm diameter) to the inner walls of polypropylene microcentrifuge tubes. In addition to increasing the surface area of the tubes manyfold, the beads provide surface Si groups which can be reacted with a silane compound such as aminopropyltriethoxysilane, yielding a free amino group. The amino group is reacted with another cross-linking reagent, for example, the homobifunctional compound dimethyl suberimidate, which can form a covalent bond with amine groups of proteins. After binding protein A or G to the dimethyl suberimidate, the beads were used to immunoprecipitate proteins from cell extracts; we show that the protein A/G-coated glass beads yield similar amounts of immunoprecipitated proteins as a standard method using protein A– or G–agarose beads, but with fewer contaminating proteins. In addition, we show that when immunoprecipitating Ras from cell extracts and measuring the amounts of Ras-bound GTP and GDP, the new method yielded higher guanine nucleotide levels than protein G–agarose beads, suggesting that it caused less denaturation of Ras. Because the glass beads are bonded to the walls of the tubes, the immunoprecipitates can be washed rapidly and efficiently, and we show that 20–30 tubes can be washed in 1/10 the time required to wash immunoprecipitates on protein A– or G–agarose beads.

The Effects of Structure on Gene Delivery with Linear β- and γ-Cyclodextrin-Containing Polycations

Cyclodextrin (CD)-containing polycations are prepared bycopolymerization of3A3B-dideoxy-3A,3B-diamino-β- and γ-CDs with dimethyl suberimidate·2HCl to yieldpolyamidine products. Both alkyl- and alkoxy-diamines are used to vary the spacing between the CDand the amidine charge centers. It is found that the transfection efficiency andtoxicity of such polycations is dramatically affected by the structure of the spacer separatingthe CD ring from the charge centers and less so by the type of CD used.

Evidence for a dimeric structure of rat liver serine dehydratase

Rat liver serine dehydratase (SDH) is known to be involved in gluconeogenesis. It has long been believed to be a dimeric protein with the subunit molecular weight ( M r) of 34,000. Recently, sheep liver SDH was reported to be a monomer with a M r of 38,000. The native M r of rat SDH was only determined by the ultracentrifugation method more than three decades ago, and that of sheep SDH was done by the method of gel chromatography. The primary to quaternary structures of a given enzyme in a specific mammalian organ are usually conserved among various species. The aim of the present investigation is to clarify the structural differences between rat and sheep SDHs. First, we found that the amino acid composition reported for sheep SDH was statistically similar to that of rat SDH. Second, immunoblot analysis using anti-rat SDH IgG as the probe showed the size of sheep SDH to be a M r of 30,500, whereas that of SDH was about M r of 35,000. On the other hand, the native size of rat SDH was assessed by two methods: (1) the laser light scattering method demonstrated that rat SDH had a M r of 66,800, consistent with the previous value ( M r=64,000); (2) cross-linking experiments of the purified rat SDH with dimethyl suberimidate revealed the existence of a dimeric form by sodium dodecyl sulfate-polyacrylamide gel electrophoresis. The present results clearly confirm that rat SDH is a dimer, and suggest that sheep SDH is similar to rat SDH immunologically, but with a molecular weight 7500 smaller than reported previously.

Dielectric spectroscopy of chemically stabilized pericardium tissue at sub-zero temperatures

In this paper dielectric spectroscopy is used as a tool for the investigation of crosslinking processes in chemically modified collageneous tissues. Measurements in the frequency range of 10/sup -1/ to 10/sup 7/ Hz and at temperatures 180, 200 and 220 K have been performed for collageneous porcine pericardium tissues, crosslinked with glutaraldehyde (GA) and dimethyl suberimidate (DMS) reagents for exposure times ranging from 15 min to 2 h. The dielectric loss spectra allowed detection of a shift of the main relaxation process to lower frequencies upon GA or DMS treatment. The observed increase of relaxation times can be interpreted as a result of the crosslinking-induced changes in dipolar polarization of collagen bound water as well as in water-assisted polarization of polar sidegroups in collagen polypeptide chains. It has been found that the relaxation time reaches a maximum value after 1 h of chemical treatment for both reagents. This result can support earlier findings, which have shown that the major part of crosslinks in collagen is created over the first hour of the chemical modification.

Changes in interfacial behaviour, emulsifying and foaming properties of faba bean legumin after modification with dimethylsuberimidate

The effect of a rising rigidity and surface hydrophobicity of the 11S storage protein from faba beans--legumin--induced by chemical modification with dimethylsuberimidate (DMS) on some surface functional properties was studied. Short-time adsorption kinetics using a droplet-volume tensiometer, pressure transformation and desorption behaviour of monolayer using a film balance, and emulsifying and foaming properties were determined to characterize surface activity and interfacial film forming behaviour. Tensio-active properties at the air-water interface, i.e. decay in surface tension and pressure transformation in monolayer, were improved by modification. However, a decrease in emulsifying activity, foam capacity and foam expansion after modification of the legumin points to an overall deterioration of energy-induced film forming behaviour. The results support the view that surface activity is generally governed more by molecular flexibility than by surface hydrophobicity.

Dimethyl suberimidate cross-linking of oligo(dT) to DNA-binding proteins.

Dimethyl suberimidate is a bifunctional reagent that is used for cross-linking the protein components of oligomeric macromolecules. In this report, dimethyl suberimidate is shown to specifically cross-link oligo(dT) of varying lengths to the DNA-binding subunits of a multimeric helicase-primase encoded by herpes simplex virus type 1. This result indicates that dimethyl suberimidate and other imidoester cross-linking reagents may be useful for characterizing the interaction of oligo(dT) with proteins that bind single-stranded DNA.