Fish-Derived Antifreeze Proteins and Antifreeze Glycoprotein Exhibit a Different Ice-Binding Property with Increasing Concentration

The concentration of a protein is highly related to its biochemical properties, and is a key determinant for its biotechnological applications. Antifreeze proteins (AFPs) and antifreeze glycoproteins (AFGPs) are structurally diverse macromolecules that are capable of binding to embryonic ice crystals below 0 °C, making them useful as protectants of ice-block formation. In this study, we examined the maximal solubility of native AFP I–III and AFGP with distilled water, and evaluated concentration dependence of their ice-binding property. Approximately 400 mg/mL (AFP I), 200 mg/mL (AFP II), 100 mg/mL (AFP III), and >1800 mg/mL (AFGP) of the maximal solubility were estimated, and among them AFGP’s solubility is much higher compared with that of ordinary proteins, such as serum albumin (~500 mg/mL). The samples also exhibited unexpectedly high thermal hysteresis values (2–3 °C) at 50–200 mg/mL. Furthermore, the analysis of fluorescence-based ice plane affinity showed that AFP II binds to multiple ice planes in a concentration-dependent manner, for which an oligomerization mechanism was hypothesized. The difference of concentration dependence between AFPs and AFGPs may provide a new clue to help us understand the ice-binding function of these proteins.

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L-Lysine: A Modulator for the Relative Activity of High Molecular Weight (HMW) and Low Molecular Weight(LHW) Urokinase(UK)

10.1055/s-0039-1687222 ◽

1979 ◽

Author(s):

L.L. Shen ◽

W.H. Holleman

Keyword(s):

Molecular Weight ◽

Caproic Acid ◽

Fibrin Clot ◽

Relative Activity ◽

Test Tube ◽

Clot Lysis ◽

Dependent Manner ◽

Plasma Clot ◽

Concentration Dependent Manner ◽

The Difference

L-Lysine(Lys), in a concentration dependent manner, progressively inhibited UK-activated lysis of human plasma clots as demonstrated by Ploug test-tube method and elastometric measurements. Lys was more effective with HMW UK than LMW UK, and the effect of Lys with LMW UK from tissue culture and urine sources was the same. Epsilon amino caproic acid(EACA) and tranexamic acid(TXA) were stronger inhibitors but inhibited HMW and LMW UK-induced lysis to the same degree. Elastometric measurements showed that Lys inhibition was not due to its interference with the initial clotting process nor to the reduction of clot rigidity. Amidolytic assays using chromogenic substrates showed that Lys had no direct effect, on UK, and that Lys enhanced the activation of the native Glu-plasminogen(Pg) by LMW UK, but not the activation by HMW UK. When the substrate was human fibrin clots, Lys enhanced the lysis induced by LMW UK while the lysis induced by HMW UK was inhibited; however, the extent of enhancement and inhibition was limited. We concluded that the mode of Lys action is not identical to that of EACA or TXA, and that the stronger Lys inhibition of plasma clot lysis as compared to fibrin clot lysis is due to the potentiation of plasma fibrinolytic inhibitors by Lys. The difference In effect of Lys on HMW and LMW UK-induced lyels is likely due to a partial conformation change of Glu-Pg molecule upon Lys binding. The relatively moderate interaction of Lys with Glu-Fg results In a mildly modified UK substrate which reacts preferentially with the enzyme smaller in size.

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Characterization of Genipin-Modified Dentin Collagen

BioMed Research International ◽

10.1155/2014/702821 ◽

2014 ◽

Vol 2014 ◽

pp. 1-7 ◽

Cited By ~ 7

Author(s):

Hiroko Nagaoka ◽

Hideaki Nagaoka ◽

Ricardo Walter ◽

Lee W. Boushell ◽

Patricia A. Miguez ◽

...

Keyword(s):

Amino Acid ◽

Lysyl Oxidase ◽

Biomechanical Properties ◽

Biochemical Properties ◽

Cross Linking ◽

Dependent Manner ◽

Cross Link ◽

Concentration Dependent Manner ◽

Cross Links ◽

Dentin Collagen

Application of biomodification techniques to dentin can improve its biochemical and biomechanical properties. Several collagen cross-linking agents have been reported to strengthen the mechanical properties of dentin. However, the characteristics of collagen that has undergone agent-induced biomodification are not well understood. The objective of this study was to analyze the effects of a natural cross-linking agent, genipin (GE), on dentin discoloration, collagen stability, and changes in amino acid composition and lysyl oxidase mediated natural collagen cross-links. Dentin collagen obtained from extracted bovine teeth was treated with three different concentrations of GE (0.01%, 0.1%, and 0.5%) for several treatment times (0–24 h). Changes in biochemical properties of NaB3H4-reduced collagen were characterized by amino acid and cross-link analyses. The treatment of dentin collagen with GE resulted in a concentration- and time-dependent pigmentation and stability against bacterial collagenase. The lysyl oxidase-mediated trivalent mature cross-link, pyridinoline, showed no difference among all groups while the major divalent immature cross-link, dehydro-dihydroxylysinonorleucine/its ketoamine in collagen treated with 0.5% GE for 24 h, significantly decreased compared to control (P< 0.05). The newly formed GE-induced cross-links most likely involve lysine and hydroxylysine residues of collagen in a concentration-dependent manner. Some of these cross-links appear to be reducible and stabilized with NaB3H4.

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Comparative Intracellular (THP-1 Macrophage) and Extracellular Activities of β-Lactams, Azithromycin, Gentamicin, and Fluoroquinolones against Listeria monocytogenes at Clinically Relevant Concentrations

Antimicrobial Agents and Chemotherapy ◽

10.1128/aac.46.7.2095-2103.2002 ◽

2002 ◽

Vol 46 (7) ◽

pp. 2095-2103 ◽

Cited By ~ 78

Author(s):

Stéphane Carryn ◽

Françoise Van Bambeke ◽

Marie-Paule Mingeot-Leclercq ◽

Paul M. Tulkens

Keyword(s):

Listeria Monocytogenes ◽

Conventional Therapy ◽

The Other ◽

Intracellular Bacteria ◽

Dependent Manner ◽

Concentration Dependent Manner ◽

Cellular Accumulation ◽

Intracellular Activities ◽

The Difference ◽

In Cells

ABSTRACT The activities of ampicillin, meropenem, azithromycin, gentamicin, ciprofloxacin, and moxifloxacin against intracellular hemolysin-positive Listeria monocytogenes were measured in human THP-1 macrophages and were compared with the extracellular activities observed in broth. All extracellular concentrations were adjusted to explore ranges that are clinically achievable in human serum upon conventional therapy. In broth, ampicillin, meropenem, and azithromycin were only bacteriostatic, whereas gentamicin, ciprofloxacin, and moxifloxacin were strongly bactericidal in a concentration-dependent manner. In cells, ampicillin, meropenem, azithromycin, and ciprofloxacin were slightly bactericidal (0.3- to 0.8-log CFU reductions), moxifloxacin was strongly bactericidal (2.1-log CFU reduction), and gentamicin was virtually inactive. The difference in the efficacies of moxifloxacin and ciprofloxacin in cells did not result from a difference in levels of accumulation in cells (6.96 ± 1.05 versus 7.75 ± 1.03) and was only partially explainable by the difference in the MICs (0.58 ± 0.04 versus 1.40 ± 0.17 mg/liter). Further analysis showed that intracellular moxifloxacin expressed only approximately 1/7 of the activity demonstrated against extracellular bacteria and ciprofloxacin expressed only 1/15 of the activity demonstrated against extracellular bacteria. Gentamicin did not increase the intracellular activities of the other antibiotics tested. The data suggest (i) that moxifloxacin could be of potential interest for eradication of the intracellular forms of L. monocytogenes, (ii) that the cellular accumulation of an antibiotic is not the only determinant of its intracellular activity (for fluoroquinolones, it is actually a self-defeating process as far as activity is concerned), and (iii) that pharmacodynamics (activity-to-concentration relationships) need to be considered for the establishment of efficacy against intracellular bacteria, just as they are for the establishment of efficacy against extracellular infections.

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Isolation and characterization of vascular smooth muscle inositol 1,4,5-trisphosphate receptor

Biochemical Journal ◽

10.1042/bj3160295 ◽

1996 ◽

Vol 316 (1) ◽

pp. 295-302 ◽

Cited By ~ 31

Author(s):

Md. Omedul ISLAM ◽

Yutaka YOSHIDA ◽

Takaki KOGA ◽

Masayasu KOJIMA ◽

Kenji KANGAWA ◽

...

Keyword(s):

Smooth Muscle ◽

Biochemical Properties ◽

Pcr Analysis ◽

Binding Property ◽

Dependent Manner ◽

Calmodulin Binding ◽

Isolation And Characterization ◽

Inositol 1,4,5 Trisphosphate ◽

Insp3 Receptor

myo-Inositol 1,4,5-trisphosphate (InsP3) receptor of porcine aorta was purified to near homogeneity and its biochemical properties were compared with those of cerebellar InsP3 receptor of the same animal species. The aortic InsP3 receptor consisted of equal amounts of two polypeptides with slightly differing molecular masses of around 240 kDa and was found to possess a single population of InsP3-binding site (Kd of 1.2 nM). The InsP3 receptor purified from porcine cerebellum was also comprised of two polypeptides. However, the molecular mass was slightly but definitely larger, being 250 kDa, and the amounts of the two polypeptides were not equal. The aortic InsP3 receptor cross-reacted with polyclonal antibody specific to type 1 InsP3 receptor as did the cerebellar InsP3 receptor. The aortic InsP3 receptor bound to calmodulin–Sepharose in a Ca2+-dependent manner, while the cerebellar InsP3 receptor did not. Reverse transcriptase-PCR analysis revealed two splicing variants of the type 1 InsP3 receptor in porcine aortic smooth muscle distinct from those of the type 1 InsP3 receptor of porcine cerebellum. The possible relevance of this difference to difference in calmodulin-binding property was discussed.

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Susceptibility of Chlamydia trachomatis to the Excipient Hydroxyethyl Cellulose: pH and Concentration Dependence of Antimicrobial Activity

Antimicrobial Agents and Chemotherapy ◽

10.1128/aac.00785-07 ◽

2008 ◽

Vol 52 (7) ◽

pp. 2660-2662 ◽

Cited By ~ 3

Author(s):

Ali A. Abdul Sater ◽

David M. Ojcius ◽

Matthew P. Meyer

Keyword(s):

Antimicrobial Activity ◽

Chlamydia Trachomatis ◽

Epithelial Cells ◽

Concentration Dependence ◽

Hydroxyethyl Cellulose ◽

Dependent Manner ◽

Concentration Dependent Manner ◽

Sexually Transmitted ◽

Calcium Sequestration ◽

Ph Dependent

ABSTRACT Hydroxyethyl cellulose (HEC) is used as a neutral excipient in microbicides used against sexually transmitted pathogens. However, HEC inhibits the infection of cervical epithelial cells by Chlamydia trachomatis at pH 5 in a concentration-dependent manner. At pH 7, infection is inversely dependent on the concentration of HEC, possibly due to pH-dependent calcium sequestration.

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An insect antifreeze protein from Anatolica polita enhances the cryoprotection of Xenopus laevis eggs and embryos

Journal of Experimental Biology ◽

10.1242/jeb.243662 ◽

2022 ◽

Author(s):

Predrag Jevtić ◽

K. Wade Elliott ◽

Shelby E. Watkins ◽

Jonathan A. Sreter ◽

Katarina Jovic ◽

...

Keyword(s):

Xenopus Laevis ◽

Antifreeze Proteins ◽

Antifreeze Protein ◽

Dependent Manner ◽

Concentration Dependent Manner ◽

Freeze Resistance ◽

Freeze Thaw ◽

Cryoprotective Agents ◽

Thaw Cycle ◽

Freeze Thaw Cycle

Cryoprotection is of interest in many fields of research, necessitating a greater understanding of different cryoprotective agents. Antifreeze proteins have been identified that have the ability to confer cryoprotection in certain organisms. Antifreeze proteins are an evolutionary adaptation that contributes to the freeze resistance of certain fish, insects, bacteria, and plants. These proteins adsorb to an ice crystal's surface and restrict its growth within a certain temperature range. We investigated the ability of an antifreeze protein from the desert beetle Anatolica polita, ApAFP752, to confer cryoprotection in the frog Xenopus laevis. X. laevis eggs and embryos microinjected with ApAFP752 exhibited reduced damage and increased survival after a freeze/thaw cycle in a concentration-dependent manner. We also demonstrate that ApAFP752 localizes to the plasma membrane in eggs and embryonic blastomeres and is not toxic for early development. These studies show the potential of an insect antifreeze protein to confer cryoprotection in amphibian eggs and embryos.

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In vitroandin vivoactivities of a bi-aryl oxazolidinone RBx 11760 against Gram positive bacteria

Antimicrobial Agents and Chemotherapy ◽

10.1128/aac.00453-16 ◽

2016 ◽

pp. AAC.00453-16 ◽

Cited By ~ 4

Author(s):

Tarani Kanta Barman ◽

Manoj Kumar ◽

Tarun Mathur ◽

Tridib Chaira ◽

G. Ramkumar ◽

...

Keyword(s):

Staphylococcus Epidermidis ◽

High Dose ◽

Dependent Manner ◽

Gram Positive ◽

Concentration Dependent Manner ◽

Gram Positive Bacteria ◽

Initial Control ◽

The Difference ◽

Time Kill ◽

Groin Abscess

RBx 11760, a bi-aryl oxazolidinone was investigated for antibacterial activity against Gram positive bacteria. The MIC90(mg/L) of RBx 11760 and linezolid againstStaphylococcus aureuswere: 2 and 4,Staphylococcus epidermidis: 0.5 and 2,Enterococcus: 1 and 4, respectively. Similarly againstStreptococcus pneumoniaeMIC90was: 0.5 and 2, respectively. In time-kill studies, RBx 11760, tedizolid and linezolid exhibited bacteriostatic effect exceptS. pneumoniae. RBx 11760 showed 2-log10kill at 4 X MIC while tedizolid and linezolid showed 2 log10and 1.4-log10kill at 16 X MIC, respectively against MRSA H-29. AgainstS. pneumoniae5051, RBx 11760 showed bactericidal activity with 4.6 log10kill at 4 X MIC compared to 2.42 log10and 1.95 log10kill of tedizolid and linezolid at 16 X MIC. RBx 11760 showed 3 h post antibiotic effects (PAE) at 4 mg/L against MRSA H-29 and linezolid showed same effect at 16mg/L. RBx 11760 inhibited the biofilm production against MRSE ATCC 35984 in concentration dependent manner. In foreign body model, linezolid and rifampicin resulted in no advantage over stasis, while same dose of RBx 11760 demonstrated a significant killing from initial control againstS. aureus(*p<0.05) and MRSE (**p<0.01). The difference in killing was statistically significant for the lower dose of RBx 11760 (*p<0.05) versus high dose of linezolid (nsp>0.05) in groin abscess model. In neutropenic mouse thigh infection, RBx 11760 showed stasis at 20 mg/kg whereas tedizolid showed same effect at 40 mg/kg. These data support the RBx 11760 as a promising investigational candidate.

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A Thermostable Aluminum-tolerant Protease Produced by FeatherDegrading Bacillus thuringiensis Isolated from Tea Plantation

Protein and Peptide Letters ◽

10.2174/0929866527666201103153309 ◽

2020 ◽

Vol 27 ◽

Author(s):

Tianwen Wang ◽

Chen Liang ◽

Sha Xiao ◽

Li Li ◽

Hongju Xu ◽

...

Keyword(s):

Circular Dichroism ◽

Serine Protease ◽

Organic Solvents ◽

Biochemical Properties ◽

Size Exclusion ◽

Dependent Manner ◽

Concentration Dependent Manner ◽

Tea Plantation ◽

Circular Dichroïsm ◽

Optimal Ph

Background: Proteases with keratinolytic activity are widely used in biotechnologies. The feather-degrading Bacillus thuringensis isolated from soil sample of a tea plantation produced high level of extracellular keratinase. Objective: This study aimed to analyze the properties by biochemical and enzymological methods to gain information for better utilization of the enzyme. Methods: The enzyme was purified with ion exchange and size exclusion chromatography. The substrate preference, optimal pH and temperature, and the effects of organic solvents and ions were checked. Circular dichroism was performed to compare the secondary structures of the native and apo-enzyme. Results: The enzyme worked best at 50 o C, and it was an acidic serine protease with an optimal pH of 6.2. Ions Ca2+ and Mg2+ were essential for its activity. Organic solvents and other metal ions generally deactivated the enzyme in a concentration-dependent manner. However, Mn2+ and DMSO, which were frequently reported as inhibitors of protease, could activate the enzyme at low concentration (0.01 to 2 mmol/L of Mn2+; DMSO <2%, v/v). The enzyme exhibited high resistance to Al3+, which might be explained by the soil properties of its host’s residence. Circular dichroism confirmed the contribution of ions to the structure and activity. Conclusion: The enzyme was a thermostable aluminum-tolerant serine protease with unique biochemical properties.

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The Characterisation and Quantification of Immobilised Concanavalin A on Quartz Surfaces Based on The Competitive Binding to Glucose and Fluorescent Labelled Dextran

Applied Sciences ◽

10.3390/app9020318 ◽

2019 ◽

Vol 9 (2) ◽

pp. 318 ◽

Cited By ~ 3

Author(s):

Trinh Hoang ◽

Bjørn Stokke ◽

Ulrik Hanke ◽

Agne Johannessen ◽

Erik Johannessen

Keyword(s):

Concanavalin A ◽

Binding Property ◽

Dependent Manner ◽

Con A ◽

Concentration Dependent Manner ◽

Specific Element ◽

Force Microscopy ◽

Atomic Force ◽

Dominant Structure ◽

Fluorescent Molecules

The competition between various carbohydrates in the binding to Concanavalin A (Con A) can be exploited in gravimetric microsensors that detect changes in mass or viscoelasticity as a function of glucose concentration. Such sensors are based on the immobilisation of Con A as the ligand specific element, and a successful application requires that the binding property of Con A is retained. This paper presents a simplified immobilisation procedure of Con A on a quartz surface, a common material for gravimetric microsensors. Structural assessment with atomic force microscopy confirmed that the surface was covered with a layer of macromolecules. This layer shows the presence of entities of various sizes, presumably monomers, dimers and tetramers among which dimers of the Con A are the most dominant structure. Functional assessment using fluorescent labelled dextran (FITC and Alexa 488) suggests a surface coverage ranging from 1.8 × 1011 to 2.1 × 1012 immobilised fluorescent molecules per cm2. The assay was responsive to glucose over a concentration range from 0–40 mM, but became gradually saturated above 20 mM. Hence, the immobilised Con A is able to bind dextran, which is displaced by glucose in a concentration dependent manner, thus triggering a mass change proportional to the MW of dextran.

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Inhibition of Streptococcus pneumoniae autolysins highlight distinct differences between chemical and genetic inactivation

10.1101/2020.09.16.300541 ◽

2020 ◽

Author(s):

Brad A Haubrich ◽

Saman Nayyab ◽

Caroline Williams ◽

Andrew Whitman ◽

Tahl Zimmerman ◽

...

Keyword(s):

Streptococcus Pneumoniae ◽

Genetic Screening ◽

Dependent Manner ◽

Cell Wall Metabolism ◽

Phenotypic Analysis ◽

Concentration Dependent Manner ◽

Chemical Inactivation ◽

Autolytic Activity ◽

The Difference ◽

Induced Autolysis

AbstractDespite renewed interest, development of chemical biology methods to study peptidoglycan metabolism has lagged in comparison to the glycobiology field in general. To address this, a panel of diamides were screened against the Gram-positive pathogen Streptococcus pneumoniae to identify inhibitors of bacterial growth. The screen identified the diamide fgkc as a narrow spectrum bacteriostatic inhibitor of S. pneumoniae growth with an MIC of 7.8 μM. The diamide inhibited detergent-induced autolysis in a concentration dependent manner indicating peptidoglycan degradation as the mode-of-action. Genetic screening of autolysin mutants suggested LytB, an endo-N-acetylglucosaminidase, involved in cell division as the potential target. Surprisingly, biochemical, and phenotypic analysis contradicted the genetic screen results. Phenotypic studies with the Δlytb strain illustrate the difference between genetic and chemical inactivation of autolysins. These findings suggest that meta-phenotypes including autolytic activity, cell morphology, and genetic screening can be the result of the complex interaction of one or more possible pathways that are connected to cell wall metabolism.

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