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At least 19 recordsLinked to original sources
PubMed1995

Biotinylation of histones by human serum biotinidase: assessment of biotinyl-transferase activity in sera from normal individuals and children with biotinidase deficiency.

Serum biotinidase has biotinyl-transferase activity in addition to biocytin hydrolase activity. A sensitive assay for biotinyl-transferase activity was developed based on the transfer of biotin from biocytin to histones. Biotinidase biotinyl-transferase occurs at physiological and alkaline pHs, whereas hydrolysis of biocytin occurs optimally at pH 4.5 to 6.0. Measurement of hydrolysis requires micromolar concentrations of biocytin, whereas biotinylation of histones can be detected readily at 1.5 nM biocytin. Because polylysine is readily biotinylated by biotinidase in the presence of biocytin, whereas polyarginine is not, the enzyme likely transfers biotin to the epsilon-amino group of lysyl residues. To determine if patients who are deficient in biocytin hydrolase activity are also deficient in biotinyl-transferase activity, serum from 103 children (25 identified by exhibiting clinical symptoms and 78 detected by newborn screening) with profound biotinidase deficiency (less than 10% of mean normal biotinyl-p-aminobenzoate hydrolyzing activity) were assessed for biotinyl-transferase activity and for the presence of cross-reacting material (CRM) to antibodies prepared against purified serum biotinidase. Sera from all symptomatic patients, both CRM-negative and CRM-positive, had no biotinyl-transferase activity. Sera that was CRM-negative from children ascertained by newborn screening also had no biotinyl-transferase activity, whereas sera from 67% of the CRM-positive children identified by newborn screening had varying degrees of biotinyl-transferase activity. These results indicate that there is a large group of enzyme-deficient children detected by newborn screening who are different biochemically from those who are symptomatic. The clinical relevance of having some degree of biotinyl-transferase activity for individuals with biotinidase deficiency remains to be determined. In addition, it is important to determine if biotinyl-transferase activity, especially biotinylation of histones, is a physiological function of biotinidase.

Amidohydrolases↗
PubMed2000

Heteronuclear NMR studies of the specificity of the post-translational modification of biotinyl domains by biotinyl protein ligase.

The lipoyl domains of 2-oxo acid dehydrogenase multienzyme complexes and the biotinyl domains of biotin-dependent enzymes have homologous structures, but the target lysine residue in each domain is correctly selected for posttranslational modification by lipoyl protein ligase and biotinyl protein ligase, respectively. We have applied two-dimensional heteronuclear NMR spectroscopy to investigate the interaction between the apo form of the biotinyl domain of the biotin carboxyl carrier protein of acetyl-CoA carboxylase and the biotinyl protein ligase (BPL) from Escherichia coli. Heteronuclear multiple quantum coherence NMR spectra of the 15N-labelled biotinyl domain were recorded in the presence and absence of the ligase and backbone amide 1H and 15N chemical shifts were evaluated. Small, but significant, changes in chemical shift were found in two regions, including the tight beta-turn that houses the lysine residue targetted for biotinylation, and the beta-strand 2 and the loop that precedes it in the domain. When compared with the three-dimensional structure, sequence alignments of other biotinyl and lipoyl domains, and mutagenesis data, these results give a clear indication of how the biotinyl domain is both recognised by BPL and distinguished from the structurally related lipoyl domain to ensure correct posttranslational modification.

Acetyl-CoA Carboxylase↗
PubMed2001

A comparison of anti-biotin and biotinylated anti-avidin double-bridge and biotinylated tyramide immunohistochemical amplification.

Often it is difficult to detect very small amounts of antigen with conventional immunohistochemical techniques. We evaluate three amplification techniques involving anti-biotin or anti-avidin double-bridges or biotinylated tyramide amplification to enhance the sensitivity of serotonin transporter immunohistochemistry. For the anti-biotin double-bridge, after the secondary antibody, the sections were incubated in anti-biotin antibody followed by a second incubation in the secondary antibody and then avidin-biotin-peroxidase complex (ABC). For the biotinylated anti-avidin technique, after the ABC, sections were incubated in biotinylated anti-avidin, followed by another incubation in ABC. For the biotinylated tyramide technique, after the ABC step, sections were incubated in biotinylated tyramide and hydrogen peroxide, followed by another incubation in ABC. The anti-biotin double-bridge also resulted in a large increase in the number of stained fibers and the intensity of labeling with no increase in background. A biotinylated anti-avidin double-bridge also produced significant signal amplification but significant background. The biotinylated tyramide technique resulted in an even larger increase in the number of labeled fibers and an intensity of their staining with a moderate amount of background staining. However, this advantage was not present at high dilutions of primary antibody. Thus, the anti-biotin double-bridge is likely to be useful in immunohistochemistry and immunofluorescence as well as other situations where increased sensitivity and low background from biotin markers is needed. The biotinylated tyramide technique may also be useful where some degree of background labeling is acceptable.

Animals↗
PubMed1996

Covalent modification of protein kinase C isozymes by the inactivating peptide substrate analog N-biotinyl-Arg-Arg-Arg-Cys-Leu-Arg-Arg-Leu. Evidence that the biotinylated peptide is an active-site affinity label.

We recently reported that the peptide substrate analog Arg-Lys-Arg-Cys-Leu-Arg-Arg-Leu (RKRCLRRL) irreversibly inactivates the protein kinase C (PKC) isozymes alpha, beta, and gamma in a dithiothreitol-sensitive manner by an active site-directed mechanism. We hypothesized that the inactivation mechanism entailed covalent complex formation between the PKC isozyme and the inactivator peptide. In this report, N-biotinylated analogs of RKRCLRRL that inactivate Ca2+-dependent PKC activity were designed and tested for their ability to covalently label PKC isozymes. A purified PKC isozyme mixture (alpha, beta, gamma, epsilon, zeta) was incubated with the N-biotinylated peptides and then subjected to denaturing gel electrophoresis, transferred to nitrocellulose, and probed for avidin-reactive species. The Ca2+-dependent PKC subfamily members PKC-alpha, -beta, and -gamma comigrated at 82 kDa and were distinguished by isozyme-specific immunoprecipitation. N-Biotinyl-RRRCLRRL covalently labeled all of the isozymes examined. When the isozymes were denatured prior to incubation with the N-biotinylated peptides, no labeling was observed. Inactivation of the Ca2+-dependent PKC subfamily by the N-biotinylated peptides was associated with covalent labeling of the 82-kDa PKC subspecies. The concentration dependence curves observed with N-biotinyl-RRRCLRRL were similar for inactivation and covalent labeling. The rank order of potency of three N-biotinylated peptides was the same for the inactivation and covalent labeling. Both the inactivation and covalent labeling were dithiothreitol-sensitive, and they were each subject to protection by MgATP and a peptide substrate analog. The covalent label was mapped to the catalytic domain of PKC by limited proteolysis of the modified enzyme. These results provide evidence that the N-biotinylated inactivator peptides are active-site affinity labels of PKC. The inactivator peptides most likely function by S-thiolating the active-site Cys residue conserved in PKC. This is the first report to demonstrate covalent labeling of PKC by a peptide substrate analog.

Animals↗
PubMed1995

Further studies on targeted DNA transfer to cells using a highly efficient delivery system of biotinylated transferrin and biotinylated polylysine complexed to streptavidin.

Conjugates consisting of biotinylated transferrin and biotinylated poly-L-lysine attached to streptavidin have been prepared and found to transfer luciferase plasmid DNA very efficiently to HeLa cells in the presence of chloroquine. Transfection was dependent on (i) use of biotinylated short chain polylysine containing 70 lysine residues, (ii) biotinylated transferrin containing 1-2 biotin moieties, (iii) reaction of biotinylated transferrin with streptavidin followed by isolation of the resulting conjugate on Sephadex G-200 and (iv) interaction of streptavidin-biotinylated transferrin with biotinylated polylysine giving a complex suitable for DNA transfection. It was found that if the above sequence of steps resulting in the formation of streptavidin-biotinylated transferrin/biotinylated polylysine was followed without isolation of intermediate conjugates by Sephadex G-200 chromatography, pRSVL DNA transfer was still very efficient. Transfer of luciferase DNA by the streptavidin conjugates and subsequent expression of luciferase activity was almost completely inhibited by excess free transferrin, showing that gene transfer was through the transferrin receptor pathway via receptor-mediated endocytosis. The streptavidin (bio2-transferrin) bio10-pLys70 conjugate used in the present experiments was approximately one hundred times more efficient in pRSVL DNA transfection with the HeLa cells than the previously described avidin-pLys460 (bio-transferrin) complex.

Bacterial Proteins↗
PubMed1992

Avidin-induced lysis of biotinylated erythrocytes by homologous complement via the alternative pathway depends on avidin's ability of multipoint binding with biotinylated membrane.

It was reported that avidin and streptavidin induce lysis of prebiotinylated red blood cells via the alternative pathway of both homologous and heterologous complement. Both of these proteins have four biotin-binding sites, providing a polyvalent interaction with biotinylated components of the erythrocyte membrane. We have compared the effects of mono- and multipoint avidin attachment on the sensitivity of biotinylated erythrocytes to lysis by the complement system. In the presence of anti-avidin antibody, avidin-bearing biotinylated erythrocytes were rapidly lysed by heterologous serum. This lysis was independent from the mode of avidin attachment, implying that complement activation by the classical pathway triggered by interaction between C1 and avidin-bound antibody on the erythrocyte surface is independent from the avidin's ability of polyvalent (multipoint) binding with biotinylated membrane components. In the absence of anti-avidin antibody, biotinylated erythrocytes bearing polyvalently attached avidin were lysed by homologous complement better than cells bearing avidin, which possesses reduced ability for multipoint binding with biotinylated erythrocyte. Two independent approaches to reduce avidin's ability of multipoint binding were used: decrease in surface density of biotin on the erythrocyte membrane and blockage of biotin-binding sites of avidin. Both methods result in reduced lysis of avidin-bearing erythrocytes as compared with erythrocytes bearing an equal amount of polyvalent-bound avidin. Thus the activation of homologous complement via the alternative pathway depends on avidin's ability to 'cross-link' to the biotinylated components of the erythrocyte membrane.

Animals↗
PubMed2002

Hepatitis C virus NS3 and simian virus 40 T antigen helicases displace streptavidin from 5'-biotinylated oligonucleotides but not from 3'-biotinylated oligonucleotides: evidence for directional bias in translocation on single-stranded DNA.

Helicases are enzymes that use energy from nucleoside triphosphate hydrolysis to unwind double-stranded (ds) DNA, a process vital to virtually every phase of DNA metabolism. Helicases have been classified as either 5'-to-3' or 3'-to-5' on the basis of their ability to unwind duplex DNA adjacent to either a 5' or 3' single-stranded (ss) DNA overhang. However, there has been debate as to whether this substrate preference is indicative of unidirectional translocation on ssDNA. We developed an assay that monitors the ability of a helicase to displace streptavidin from biotinylated oligonucleotides [Morris, P. D., and Raney, K. D. (1999) Biochemistry 38, 5164-5171]. Two helicases identified as having 5'-to-3' polarity displaced streptavidin from the 3'-end of biotinylated oligonucleotides but not from the 5'-end. We performed similar experiments using the 3'-to-5' helicases from the hepatitis C virus (NS3) and SV40 virus (SV40 T antigen). NS3 and SV40 T antigen were able to displace streptavidin from a 5'-biotinylated oligonucleotide but not from a 3'-biotinylated oligonucleotide. NS3 and SV40 T antigen enhanced the spontaneous rate of dissociation of streptavidin from biotin 340-fold and 1700-fold, respectively. The ssDNA binding protein, gp32, did not enhance dissociation of streptavidin from either end of an oligonucleotide. For NS3, the rate of displacement was faster from a 5'-biotinylated 60mer than from a 5'-biotinylated 30mer. The strong directional bias in streptavidin displacement activity exhibited by each helicase is consistent with a directional bias in translocation on ssDNA. The dependence of the reaction with NS3 on the oligonucleotide length suggests that multiple NS3 monomers are necessary for optimal activity.

Antigens, Polyomavirus Transforming↗
PubMed1992

Cytotoxicity of streptavidin-blocked biotinyl-ricin is retrieved by in vitro immunotargeting via biotinyl monoclonal antibody.

The streptavidin-biotin system has been used to immunotarget whole ricin to tumor cells in a system that overcomes ricin-nonspecific cytotoxicity. Biotin was linked to ricin via a disulfide-containing reagent, sulfosuccinimidyl-2-(biotinamido)ethyl-1,3'-dithiopropionate. The product, biotinyl-S,S-ricin (b-ricin), retained most of its in vitro cytotoxic activity against human epidermoid carcinoma (KB) cells. Complexing b-ricin to streptavidin resulted in greater than 99% loss of its cellular toxicity which is associated with loss of cell-binding activity. The streptavidin-b-ricin complex could, however, be targeted to KB cells via the biotinylated monoclonal antibody 108 which is specific to the epidermal growth factor receptor overexpressed on KB cells. The complex did not regain its activity if the specific antibody was not biotinylated or if the biotinylated antibody was of a different specificity. Streptavidin is thus used to block b-ricin, presumably due to a steric restraint of the streptavidin on the ricin B-chain, and to bridge it to biotinyl antibody recognizing the target cell. Avidin could not replace streptavidin in this system since a complex between b-ricin and avidin retained a major part (60%) of ricin cytotoxic activity. This is attributed to the nonspecific binding of avidin to cells in vitro, including the KB cells. It is suggested that b-ricin is blocked by both streptavidin and avidin, but once the complex gains access to the cell surface, its cytotoxic activity is specifically retrieved.

Avidin↗
PubMed1988

Reversible biotinylation of C1q with a cleavable biotinyl derivative. Application in C1q receptor (C1qR) purification.

Reversible biotinylation of human C1q without impairment of its physiologic functions has allowed us to develop a simple and rapid purification method for C1q receptor (C1qR). The biotinylating reagent, NHS-SS-biotin (Mr 606.7) contains an extended connector or cross-linker arm which limits steric hindrance and is bridged by a cleavable disulfide bond to the biotin component. Biotinylation was achieved by mixing C1q (in PBS, pH 7.4) with NHS-SS-biotin (dissolved in dimethyl formamide) in a 50:1 v/v and 1:25 mol/mol ratio and allowing the reaction to continue at room temperature for 4 h. The mixture was then dialyzed against PBS pH 7.4 (2 X 1 liter) and analyzed by SDS-PAGE and hemolytic assay using C1q depleted serum. Under these conditions neither denaturation of the protein nor loss of hemolytic activity was evident. Such biotinylated C1q (Bio-C1q) was used to pull out the C1qR from detergent-solubilized (1% NP-40 in PBS, pH 7.4 plus inhibitors) 125I-surface labeled membrane solution that had been first centrifuged (1 h, 45,000 X g, 4 degrees C) and then sequentially precleared with immobilized protein A, protein A-IgG and gelatin. The mixture of Bio-C1q and membrane solution was then incubated (20 h, 4 degrees C), applied to immobilized avidin (equilibrated with PBS, pH 7.4, 0.1% NP-40) and after washing, the bound C1qR was eluted with equilibrating buffer containing 1 M NaCl, and the C1q by same buffer containing 100 mM DTT. The eluted C1qR contained a major Mr 70,000 molecule which upon reduction electrophoresed with an apparent Mr of 85,000-90,000 as assessed by SDS-PAGE analysis. In addition, a faint single chain band of 30-40 kDa was eluted with the major band and may represent a non-covalently associated part of the C1qR molecule.

Avidin↗
PubMed1991

Biotinylated peptides/proteins. II. Identification of biotinylated lysyl phenylthiohydantoins.

The identification and characterization of biotinylated lysyl residues in a polypeptide chain by automated sequence analysis is described. An in depth analytical study was conducted for the delineation of N epsilon modification of lysyl residues with N-hydroxysuccinimide esters of biotin and 6-aminohexanoic biotin. Confirmation of the structure of the phenylthiohydantoin derivatives of N epsilon biotinylated lysine was achieved by mass spectrometry. The analytical study focused on the identification of biotinylated lysine-4 of neuropeptide Y which served as a model peptide for the analytical procedures detailed.

Amino Acid Sequence↗
PubMed1991

Two-dimensional crystals of streptavidin on biotinylated lipid layers and their interactions with biotinylated macromolecules.

Streptavidin forms two-dimensional crystals when specifically bound to layers of biotinylated lipids at the air/water interface. The three-dimensional structure of streptavidin determined from the crystals by electron crystallography corresponds well with the structure determined by x-ray crystallography. Comparison of the electron and x-ray crystallographic structures reveals the occurrence of free biotin-binding sites on the surface of the two-dimensional crystals facing the aqueous solution. The free biotin-binding sites could be specifically labeled with biotinylated ferritin. The streptavidin/biotinylated lipid system may provide a general approach for the formation of two-dimensional crystals of biotinylated macromolecules.

Bacterial Proteins↗
PubMed1989

[Inclusion of biotinylated analogs of dUTP and dCTP in DNA by DNA-polymerases. Cloning DNA fragments, containing biotinylated deoxyribouridine in E. coli].

The synthesis of a biotinylated derivative of dCTP, viz. N4-[(N-biotinyl)-4-amino-butoxyl]-2'-deoxycytidine 5'-triphosphate (I), is described. DNA polymerase I (Klenow fragment) incorporates (I) in DNA chains instead of thymidine, although with a lower efficiency than previously described biotinylated dUTP derivative (II), whereas highly purified DNA polymerase alpha from human placenta uses as substrate derivative (II) but not (I). A DNA fragment bearing biotin residues in one of strands was synthesized with the use of DNA polymerase alpha and dUTP derivative (II); its cloning in the plasmid vector pBR322 revealed that the DNA nucleotide sequence remained intact.

Base Sequence↗
PubMed2002

Visualisation of hyaluronan and hyaluronan-binding proteins within ovine vertebral cartilages using biotinylated aggrecan G1-link complex and biotinylated hyaluronan oligosaccharides.

The aim of this study was to localise hyaluronan (HA)-binding proteins (HABPs) in ovine vertebral tissues using biotinylated HA oligosaccharides (bHA oligos) as novel affinity probes and to compare this with the distribution of tissue HA visualised using biotinylated aggrecan G1 domain-link protein complex. The bHA oligos, with a size of 6-18 disaccharides were prepared by partial digestion of HA with ovine testicular hyaluronidase, labelled with biotin hydrazide and purified by a combination of aggrecan G1 domain and avidin affinity chromatography. Hyaluronan and HABPs were both prominent pericellular components of hypertrophic cells of the vertebral epiphyseal growth plate and enlarged cells in the cartilaginous end plate of the disc. The bHA oligo probe also visualised HABPs intracellularly in hypertrophic cells, which also contained intracellular HA. Monolayer cultures of ovine annulus fibrosus and nucleus pulposus cells rapidly internalised the bHA oligo affinity probe which was subsequently visualised by indirect fluorescence using avidin-FITC, to cytoplasm and discrete nuclear regions. The results indicate that the abundant pericellular and intracellular HA associated with cartilaginous cells in the vertebral tissues is colocalised with HABPs. The bHA oligo affinity probe may have further applications in investigations of intracellular HABPs, HA endocytosis and the roles they play in cellular regulatory processes.

Aggrecans↗
PubMed1987

The effect of pH on the aggregation of biotinylated antibodies and on the signal-to-noise observed in immunoassays utilizing biotinylated antibodies.

During the development of immunoassays to detect gram-negative bacteria, an effect of pH on the aggregation of some murine monoclonal antibodies directed to Neisseria gonorrhoeae was observed. By reacting positively charged primary amines on these antibodies with the neutral NHS-biotin (N-hydroxy-succinimidobiotin), the surface charge on the antibodies was altered and a concomitant change in the solubility of these antibodies noted. This derivatization produced not only a pH-dependent change in the solubility properties of the antibodies, but also affected the response of immunoassays in which these antibodies were used. Data presented suggests that the signal-to-noise (S/N) observed in these assays is maximized under conditions where the biotinylated antibody is introduced into the assay at a pH at least 2 U above its pI. Our hypothesis is that as the pH of the solution approaches the biotinylated antibodies' isoelectric point, they become 'stickier', perhaps by aggregation (which we have directly measured), leading to high non-specific binding and hence a lower S/N.

Antibodies, Bacterial↗
PubMedDate not supplied

A stable bis-allyloxycarbonyl biotin aldehyde derivative for biotinylation via reductive alkylation: application to the synthesis of a biotinylated doxorubicin derivative.

A novel, stable, biotin aldehyde derivative is reported in which the biotin moiety is N1,N3-protected by the allyloxycarbonyl group. The derivative is stable to sodium cyanoborohydride mediated reductive alkylation and is cleaved under mild Pd [0] catalysis. This novel biotin aldehyde should have wide application in avidin- and streptavidin-based detection systems and bioassays. The derivative is utilized in the synthesis of a biotinylated doxorubicin analogue that retains topoisomerase activity.

Affinity Labels↗
PubMedDate not supplied

A novel intravascular drug delivery method using endothelial biotinylation and avidin-biotin binding.

In this study, a novel intravascular drug delivery system was developed in which a drug injected from a catheter was fixed to the vasculature of the targeted tissue. Cellular proteins of viable endothelial cells were first biotinylated directly by biotinylation reagents, and then bound by an avidinated drug or, using avidin as a linker, a biotinylated drug. In the initial experiments, we studied in vitro the biotinylation of cultured bovine aortic endothelial cells (BAECs) by applying biotinylation reagents (NHS-LC-biotin or sulfo-NHS-LC-biotin) onto the washed intact BAEC monolayers and showed that the amount of biotin bound to the cells depended on the concentration of the biotinylation reagents applied. The cell-bound biotin decreased with time after the biotinylation. When fluorescein-labeled avidin (FITC-avidin) was applied to the biotinylated BAEC monolayers, the FITC-avidin readily bound to the cells. An LDH-release assay showed that sulfo-NHS-LC-biotin was only slightly cytotoxic to the BAECs and a colony formation assay showed only slight adverse effects of the reagent. In vivo studies were carried out on the renal arteries of normal rabbits. A solution of NHS-LC-biotin was injected through a catheter to one kidney to biotinylate its vasculature and the vehicle to the other as control, followed by a perfusion with saline. Finally, a solution of FITC-avidin was injected to both kidneys that were then reperfused with the blood flow following the withdrawal of the catheters. In the histological sections, more than 85% of glomeruli was stained with fluorescein in the biotinylated kidney, whereas no glomeruli were stained in the control. In the kidneys harvested 2 days after the same procedure, most glomeruli were still brightly stained. In the final experiment, biotinylated kidneys were injected with a solution of avidin, followed by a solution of fluorescein-biotin. Control kidneys had no prior biotinylation but received the same injections of avidin and fluorescein-biotin as above. More than 80% of glomeruli were stained in the biotinylated kidneys but none in the controls. This indicated that biotinylated drugs can be anchored to the biotinylated vasculature through avidin without being flushed away by blood flows. No apparent adverse effect was found in the functions of biotinylated kidneys. We propose that this drug delivery system is feasible for the treatment of some pathological conditions of blood vessels such as microvascular proliferation in malignant tumors and for continuous drug delivery in certain target organs.

Animals↗
PubMed1996

Biotinylation of proteins via amino groups can induce binding to U937 cells, HL-60 cells, monocytes and granulocytes.

The use of biotinylated ligands for the flow cytometric detection of cell surface receptors has become a popular alternative to radioreceptor assays. Although the biotinylation of a protein is a relatively mild chemical reaction several reports have mentioned the fact that the number and location of biotin moieties coupled to amino groups of a protein can alter its physicochemical properties and impair biological activity. In the present study we show for a variety of biotinylated functionally unaltered ligands that biotinylation by N-hydroxysuccinimide (NHS) esters of biotin can induce a binding to cell surfaces, which is not specific for the respective unlabelled ligand. C1q, C1 inhibitor (C1-INH), alpha 1-antitrypsin (AT), ovalbumin (OV), transferrin and soybean trypsin inhibitor (STI) were labelled with S-NHS-LC-biotin and activated C1s (C1s) with NHS-biotin. Biotinylation of C1q, C1s and C1-INH exerted negligible effects on biological function, antigenicity or electrophoretic mobility but when labelled and unlabelled proteins were assayed for binding to monocytic U937 cells, promyelocytic HL-60 cells, monocytes and granulocytes, a remarkable binding was observed for biotinylated C1q, C1-INH and C1s. In contrast, no binding was observed when we used unlabelled C1q, C1s and C1-INH and employed specific antibodies, alpha-mouse-FITC or alpha-rabbit-FITC for detection. Increasing molar ratios of biotin-to-protein (B : P) for biotinylated AT, OV and STI evoked increased fluorescence intensities of the cells. Most importantly the unlabelled ligands did not compete for cell binding with their biotinylated derivatives, with the exception of transferrin. Preincubation of the cells with an excess of free d-biotin did not reduce binding of biotinylated proteins, thus excluding a potential involvement of biotin receptors. Hydrophobic interaction chromatography revealed a remarkable increase in hydrophobicity of the biotinylated proteins compared to their unlabelled counterparts, suggesting that the biotinylation-induced binding is due to increased hydrophobicity. Our findings indicate that biotinylation by the common amino acid esterification method may be critical for proteins if they are to be used as ligands for receptor binding studies.

Amino Acids↗