Ice Nucleation Protein Research Articles

Bacillus Cellulase Molecular Cloning, Expression, and Surface Display on the Outer Membrane of Escherichia coli.

One of the main challenges of using recombinant enzymes is that they are derived from genetically-modified microorganisms commonly located in the intracellular region. The use of these recombinant enzymes for commercial purposes requires the additional processes of cell disruption and purification, which may result in enzyme loss, denaturation, and increased total production cost. In this study, the cellulase gene of Bacillus licheniformis ATCC 14580 was cloned, over-expressed, and surface displayed in recombinant Escherichia coli using an ice-nucleation protein (INP). INP, an outer membrane-bound protein from Pseudomonas syringae, was utilized as an anchor linker, which was cloned with a foreign cellulase gene into the pET21a vector to develop a surface display system on the outer membrane of E. coli. The resulting strain successfully revealed cellulase on the host cell surface. The over-expressed INP-cellulase fusion protein was confirmed via staining assay for determining the extracellular cellulase and Western blotting method for the molecular weight (MW) of cellulase, which was estimated to be around 61.7 kDa. Cell fractionation and localization tests demonstrated that the INP-cellulase fusion protein was mostly present in the supernatant (47.5%) and outer membrane (19.4%), while the wild-type strain intracellularly retained enzymes within cytosol (>61%), indicating that the INP gene directed the cellulase expression on the bacteria cell surface. Further studies of the optimal enzyme activity were observed at 60 °C and pH 7.0, and at least 75% of maximal enzyme activity was preserved at 70 °C.

Effects of Ice Nucleation Protein Repeat Number and Oligomerization Level on Ice Nucleation Activity

AbstractIce nucleation active bacteria have attracted particular attention due to their unique ability to produce specific ice nucleation proteins (INpros), which are the most efficient ice nuclei known as they induce nucleation at temperatures close to 0°C. Our model bacterium Pseudomonas syringae strain R10.79 produced INpros containing 67 tandem repeats, forming the proposed ice‐binding surface. To understand the role of the INpro repeats as well as the role of intermolecular interactions between INpros for their ice nucleation behavior, we produced a truncated version of the protein with only 16 tandem repeats (INpro16R). The purified INpro16R produced oligomers of varying sizes. Immersion freezing ice nucleation behavior of purified INpro16R was characterized by droplet‐freezing assays and in the Leipzig Aerosol Cloud Interaction Simulator. Predominant INpro16R oligomers introduced into Leipzig Aerosol Cloud Interaction Simulator as single particles with diameters of 50 nm or 70 nm were ice nucleation active at temperatures of −26°C and −24°C, respectively. These are much lower temperatures compared to that of intact INpros (−12°C). The data clearly indicated that the number of repeats determines the ice nucleation temperature. In addition, ice nucleation between −9°C and −10°C, comparable to the activity of intact INpro, was caused by higher‐order INpro16R oligomers. This supported previous observations that INpro oligomerization increases the ice‐binding surface, thereby affecting ice nucleation activity. In conclusion, both repeat number and oligomerization contribute in a seemingly independent manner to the nucleation mechanism of INpros.

Detection of catechol using an electrochemical biosensor based on engineered Escherichia coli cells that surface-display laccase

In this study, we report an electrochemical microbial biosensor that was made by immobilizing a bacterial laccase on the surface of Escherichia coli cells followed by adsorption of modified live cells onto a glassy-carbon electrode. Expression and surface localization of laccase on target cells were confirmed by Western blotting, flow cytometry assays and immunofluorescence microscopy observation. Increased tandem-aligned anchors with three repeats of the N-terminal domain of an ice nucleation protein were used to construct a highly active E. coli whole cell laccase-based catalytic system. When the proposed biosensor was used to detect catechol, the electrochemical response under optimized pH conditions was linear within a concentration range of 0.5 μM–300.0 μM catechol. Metal ions (Mn2+, Fe3+, Cu2+, Mg2+, Al3+ and Zn2+) at concentrations from 1 to 10 mg L−1, bovine serum albumin and glucose at concentrations from 0.1 to 10 g L−1, and ascorbic acid at concentrations from 0.01 to 0.1 g L−1 did not cause a noticeable interference effect. The detection limit of 0.1 μM catechol was comparable to those of other biosensors based on purified chemically modified laccases. When used to detect catechol in real red wine and tea samples, the biosensor offered a considerable level of accuracy comparable to the HPLC method as well as high recovery rates (98.2%–103.8%) towards all of the tested samples. Moreover, the developed system also exhibited high stability and reproducibility.

Improved freeze drying efficiency by ice nucleation proteins with ice morphology modification

This study aims to use ice nucleation proteins (INPs) as a novel approach to improve the efficiency of freeze drying process and investigate the related mechanism of ice morphology. Our results show that INPs can significantly improve freeze drying efficiency with increased primary drying rate under the increase of INP concentration from 0 to 10−2mg/mL. Moreover, such improvement was more significant at higher subzero freezing temperatures with the addition of INPs, when the control samples were unable to freeze. Those improvements further lead to reduced total drying time, which suggests an estimated total energy saving of 28.5% by INPs. Our ice morphology results indicate the ability of INPs to alter ice morphology with lamellar ice structure and larger crystal size, which both show linear relationships with primary drying rate. The results further suggest that these ice morphology characteristics induced by INPs are very likely to facilitate the water vapor flow and improve the sublimation rate. Additionally, the increase of freeze drying efficiency can also be achieved by INPs in other food systems like coffee and milk with elevated primary drying rate. The results of this study suggest great potential of using INPs to improve the efficiency of freeze drying process for a wide range of food products and other related applications. This study also provides new insights into the relationship between process efficiency and ice morphology.

Display of GCRV vp7 protein on the surface of Escherichia coli and its immunoprotective effects in grass carp (Ctenopharyngodon idella)

Infection with Grass carp reovirus (GCRV) is becoming unprecedentedly widespread in grass carp (Ctenopharyngodon idella) aquaculture industry, yet the management of GCRV infection still remains a challenge. Therefore, it is of importance to develop effective means against GCRV. As a delivery system of viral antigens, surface displaying of heterologous proteins on bacteria using anchoring motifs has successfully been implemented in human and veterinary vaccines research. In this study, a novel vaccine (BL21/InpN/vp7) was developed based on surface displaying a major capsid protein (vp7) of GCRV using the anchoring motif of N-terminal unique domain of ice-nucleation protein (InpN) on Escherichia coli BL21 (DE3) vaccine. Then the grass carp were immunized by surface displaying BL21/InpN/vp7 vaccine against GCRV using both intraperitoneal injection and bath immunization and their immune responses were tested. The results revealed that some non-specific immune parameters (acid phosphatase (ACP), alkaline phosphatase (AKP) and total antioxidant capacity (T-AOC)) were strongly increased in grass carp post injection inoculation (vp7 dose ranged from 10 to 20 μg). The specific antibody levels against GCRV and the transcriptional of immune-related genes (TNF-α, IL-1β, MHCI and IgM) were also significantly enhanced in grass carp by injection inoculation (vp7 dose ranged from 5 to 20 μg). On the other hand, only the highest dose of bath vaccination significantly induced the production of specific antibody and up-regulated transcriptions of several immune-related genes (IgM and MHCI) in grass carp. The lower cumulative mortality of grass carp in vaccinated groups after GCRV challenge clearly demonstrated that surface displayed vp7 vaccine could protect fish against GCRV infection. The relative percentage survival (RPS) value in injection vaccinated group (88.89%) was much higher compared to bath group (18.89%), which was in consistent with the production of specific serum antibodies, non-specific immune response and immune related genes expression. To sum up, our results indicated the surface display of heterologous antigenic proteins on E. coli BL21 (DE3) using the anchoring motif of ice-nucleation protein may provide a promising approach to the vaccine development of aquatic animals and suggested its potential to be used as vaccine to fight against GCRV infection.

Simultaneous hydrolysis of carbaryl and chlorpyrifos by Stenotrophomonas sp. strain YC-1 with surface-displayed carbaryl hydrolase

Many sites are often co-contaminated with multiple pesticides. To date, there are no reports on simultaneous degradation of different classes of pesticides by a natural microorganism. In this work, we aim at constructing a live biocatalyst able to simultaneously hydrolyze carbaryl and chlorpyrifos. For this purpose, carbaryl hydrolase (CH) was displayed on the cell surface of a chlorpyrifos-degrading bacterium Stenotrophomonas sp. strain YC-1 using N- and C-terminal domain of ice nucleation protein (INPNC) from Pseudomonas syringae INA5 as an anchoring motif. The localization of INPNC-CH fusion protein in the outer membrane fraction was demonstrated by cell fractionation followed by Western blot analysis. Surface display of INPNC-CH was further confirmed by proteinase accessibility experiment and immunofluorescence microscope. CH was present in an active form on cell surface without causing any growth inhibition, suggesting that the INP-based display system is a useful tool for surface expression of macromolecular heterologous proteins on the bacterial cell surface. Because surface-displayed CH has free access to pesticides, this bacterium can be used as a whole-cell biocatalyst for efficient hydrolysis of pesticides.

Draft Genome Sequence of Pseudomonas syringae PDD-32b-74, a Model Strain for Ice-Nucleation Studies in the Atmosphere.

ABSTRACTWe report here the whole genome sequence of Pseudomonas syringae PDD-32b-74, a gammaproteobacterium isolated from cloud water. This microorganism is equipped with ice-nucleation protein and biosurfactant genes that could potentially be involved in physicochemical processes in the atmosphere and clouds.

Use of Ice-Nucleating Proteins To Improve the Performance of Freeze-Thaw Valves in Microfluidic Devices.

Currently, reliable valving on integrated microfluidic devices fabricated from rigid materials is confined to expensive and complex methods. Freeze-thaw valves (FTVs) can provide a low cost, low complexity valving mechanism, but reliable implementation of them has been greatly hindered by the lack of ice nucleation sites within the valve body's small volume. Work to date has required very low temperatures (on the order of -40 °C or colder) to induce freezing without nucleation sites, making FTVs impractical due to instrument engineering challenges. Here, we report the use of ice-nucleating proteins (INPs) to induce ice formation at relatively warm temperatures in microfluidic devices. Microfluidic channels were filled with buffers containing femtomolar INP concentrations from Pseudomonas syringae. The channels were cooled externally with simple, small-footprint Peltier thermoelectric coolers (TECs), and the times required for channel freezing (valve closure) and thawing (valve opening) were measured. Under optimized conditions in plastic chips, INPs made sub-10 s actuations possible at TEC temperatures as warm as -13 °C. Additionally, INPs were found to have no discernible inhibitory effects in model enzyme-linked immunosorbent assays or polymerase chain reactions, indicating their compatibility with microfluidic systems that incorporate these widely used bioassays. FTVs with INPs provide a much needed reliable valving scheme for rigid plastic devices with low complexity, low cost, and no moving parts on the device or instrument. The reduction in freeze time, accessible actuation temperatures, chemical compatibility, and low complexity make the implementation of compact INP-based FTV arrays practical and attractive for the control of integrated biochemical assays.

Comparison among Japanese isolates of Pseudomonas savastanoi pv. savastanoi, causal agent of olive knot disease

Olive knot disease in Japan was first reported in Shizuoka Prefecture in 2014, and the causal agent was identified as Pseudomonas savastanoi pv. savastanoi. Subsequently, olive trees having knots were also found in Aichi and Kanagawa Prefectures in 2015, and the isolates from knots were also suspected to be P. savastanoi pv. savastanoi through preliminary examinations. Therefore, the Aichi and Kanagawa isolates were identified through comparison of isolates from three prefectures. Phylogenic analysis based on 16S rDNA and housekeeping genes (gyrB, rpoD, gltA and gap1) revealed that the isolates belonged to the same cluster as the pathotype strain, ICMP4352PT. The iaaM, H and L genes, which are involved in promotion of symptoms, and the ina gene coding the ice nucleation protein, were detected by PCR from all the isolates. In rep-PCR (ERIC and REP) analyses, the isolates yielded DNA fragment-banding patterns that were nearly identical to that of ICMP4352PT, but slight variations in banding patterns were observed among them. In a pathogenicity test, the isolates formed distinct knots on olive and pink jasmine. Phenotypic properties of the isolates were almost identical to those of ICMP4352PT, with the exception of d-sorbitol utilization. Consequently, Aichi and Kanagawa isolates from olive were identified as P. savastanoi pv. savastanoi, and several genetic diversities in terms of rep-PCR were found in the Japanese population of P. savastanoi pv. savastanoi, indicating their heterogeneity.

Display of P Domain of Human Norovirus Major Capsid Protein on Escherichia Coli Surface

Human noroviruses (HuNoVs) are the major viral pathogens of epidemic acute gastroenteritis. To develop an effective method for mining functional receptors of HuNoVs is the first step to know the interaction mechanism between HuNoVs and their hosts. A novel approach that incorporates the mining of viral receptors was introduced in this study by targeting the P domain of the major capsid protein from HuNoVs (GI.1 or GII.4) onto the surface of Escherichia colivia the N-terminal domain of the ice nucleation protein InaQ (InaQN). Results of growth curves demonstrated that the expression of fusion proteins did not influence the bacterial growth. After optimization of induction conditions, immunofluorescence microscopy was employed to confirm bacterial surface localization and antigenicity. Results revealed remarkable surface display ability and antigenicity properties. Therefore, bacterial surface displayed P domain can be a good candidate for further mining HuNoVs receptors in the environments.

A Novel Approach To Improve the Efficiency of Block Freeze Concentration Using Ice Nucleation Proteins with Altered Ice Morphology.

Freeze concentration is a separation process with high success in product quality. The remaining challenge is to achieve high efficiency with low cost. This study aims to evaluate the potential of using ice nucleation proteins (INPs) as an effective method to improve the efficiency of block freeze concentration while also exploring the related mechanism of ice morphology. Our results show that INPs are able to significantly improve the efficiency of block freeze concentration in a desalination model. Using this experimental system, we estimate that approximately 50% of the energy cost can be saved by the inclusion of INPs in desalination cycles while still meeting the EPA standard of drinking water (<500 ppm). Our investigative tools for ice morphology include optical microscopy and X-ray computed tomography imaging analysis. Their use indicates that INPs promote the development of a lamellar structured ice matrix with larger hydraulic diameters, which facilitates brine drainage and contains less brine entrapment as compared to control samples. These results suggest great potential for applying INPs to develop an energy-saving freeze concentration method via the alteration of ice morphology.

Diversity and Abundance of Ice Nucleating Strains of Pseudomonas syringae in a Freshwater Lake in Virginia, USA.

The bacterium Pseudomonas syringae is found in a variety of terrestrial and aquatic environments. Some strains of P. syringae express an ice nucleation protein (hereafter referred to as Ice+) allowing them to catalyze the heterogeneous freezing of water. Though P. syringae has been sampled intensively from freshwater sources in France, little is known about the genetic diversity of P. syringae in natural aquatic habitats in North America. We collected samples of freshwater from three different depths in Claytor Lake, Virginia, USA between November 2015 and June 2016. Samples were plated on non-selective medium (TSA) and on medium selective for Pseudomonas (KBC) and closely related species to estimate the total number of culturable bacteria and of Pseudomonas, respectively. A droplet freezing assay was used to screen colonies for the Ice+ phenotype. Ice+ colonies were then molecularly identified based on the cts (citrate synthase) gene and the 16S rDNA gene. Phylogenetic analysis of cts sequences showed a surprising diversity of phylogenetic subgroups of P. syringae. Frequencies of Ice+ isolates on P. syringae selective medium ranged from 0 to 15% per sample with the highest frequency being found in spring. Our work shows that freshwater lakes can be a significant reservoir of Ice+ P. syringae. Future work is needed to determine the contribution of P. syringae from freshwater lakes to the P. syringae populations present in the atmosphere and on plants and, in particular, if freshwater lakes could be an inoculum source of P. syringae-caused plant disease outbreaks.

Functional display of ice nucleation protein InaZ on the surface of bacterial ghosts

ABSTRACT In a concept study the ability to induce heterogeneous ice formation by Bacterial Ghosts (BGs) from Escherichia coli carrying ice nucleation protein InaZ from Pseudomonas syringae in their outer membrane was investigated by a droplet-freezing assay of ultra-pure water. As determined by the median freezing temperature and cumulative ice nucleation spectra it could be demonstrated that both the living recombinant E. coli and their corresponding BGs functionally display InaZ on their surface. Under the production conditions chosen both samples belong to type II ice-nucleation particles inducing ice formation at a temperature range of between −5.6 °C and −6.7 °C, respectively. One advantage for the application of such BGs over their living recombinant mother bacteria is that they are non-living native cell envelopes retaining the biophysical properties of ice nucleation and do no longer represent genetically modified organisms (GMOs).

Bacterial Surface-Displayed GII.4 Human Norovirus Capsid Proteins Bound to HBGA-Like Molecules in Romaine Lettuce.

Human Noroviruses (HuNoVs) are the main cause of non-bacterial gastroenteritis. Contaminated produce is a main vehicle for dissemination of HuNoVs. In this study, we used an ice nucleation protein mediated surface display system to present the protruding domain of GII.4 HuNoV capsid protein on bacterial surface and used it as a new strategy to explore interaction between HuNoV protein and receptor candidates from romaine lettuce. The surface-displayed HuNoV proteins were confirmed on the surface of the transformed bacteria by an immunofluorescence assay. The distribution patterns of the surface-displayed HuNoV proteins in romaine lettuce were identified through a confocal immunofluorescence assay. The surface-displayed HuNoV proteins could be found in the stomata, and the surfaces of vein and leaf of romaine lettuce. The surface-displayed HuNoV proteins could be captured by an ELISA assay utilizing extract from leaf (LE) or vein (VE). The binding of the surface-displayed HuNoV proteins to LE or VE could be competitively blocked by histo-blood group antigens from human saliva. In addition, the binding of the surface-displayed HuNoV proteins to LE or VE could also be attenuated by heat denaturation of lettuce proteins, and abolished by oxidation of lettuce carbohydrates. The results indicated that histo-blood group antigen-like molecules in LE or VE were involved in the binding of the surface-displayed HuNoV proteins to romaine lettuce. All data demonstrated that the surface-displayed HuNoV proteins could be utilized in a new and simple system for investigation of the interaction between the HuNoVs and their candidate ligands.

Ice and anti-nucleating activities of an ice-binding protein from the annual grass, Brachypodium distachyon.

Plants exposed to sub-zero temperatures face unique challenges that threaten their survival. The growth of ice crystals in the extracellular space can cause cellular dehydration, plasma membrane rupture and eventual cell death. Additionally, some pathogenic bacteria cause tissue damage by initiating ice crystal growth at high sub-zero temperatures through the use of ice-nucleating proteins (INPs), presumably to access nutrients from lysed cells. An annual species of brome grass, Brachypodium distachyon (Bd), produces an ice-binding protein (IBP) that shapes ice with a modest depression of the freezing point (~0.1 °C at 1 mg/mL), but high ice-recrystallization inhibition (IRI) activity, allowing ice crystals to remain small at near melting temperatures. This IBP, known as BdIRI, is unlike other characterized IBPs with a single ice-binding face, as mutational analysis indicates that BdIRI adsorbs to ice on two faces. BdIRI also dramatically attenuates the nucleation of ice by bacterial INPs (up to -2.26 °C). This 'anti-nucleating' activity is significantly higher than previously documented for any IBP.

Boreal pollen contain ice-nucleating as well as ice-binding \u2018antifreeze\u2019 polysaccharides

Ice nucleation and growth is an important and widespread environmental process. Accordingly, nature has developed means to either promote or inhibit ice crystal formation, for example ice-nucleating proteins in bacteria or ice-binding antifreeze proteins in polar fish. Recently, it was found that birch pollen release ice-nucleating macromolecules when suspended in water. Here we show that birch pollen washing water exhibits also ice-binding properties such as ice shaping and ice recrystallization inhibition, similar to antifreeze proteins. We present spectroscopic evidence that both the ice-nucleating as well as the ice-binding molecules are polysaccharides bearing carboxylate groups. The spectra suggest that both polysaccharides consist of very similar chemical moieties, but centrifugal filtration indicates differences in molecular size: ice nucleation occurs only in the supernatant of a 100 kDa filter, while ice shaping is strongly enhanced in the filtrate. This finding may suggest that the larger ice-nucleating polysaccharides consist of clusters of the smaller ice-binding polysaccharides, or that the latter are fragments of the ice-nucleating polysaccharides. Finally, similar polysaccharides released from pine and alder pollen also display both ice-nucleating as well as ice-binding ability, suggesting a common mechanism of interaction with ice among several boreal pollen with implications for atmospheric processes and antifreeze protection.

A one-step procedure for immobilising the thermostable carbonic anhydrase (SspCA) on the surface membrane of Escherichia coli

The carbonic anhydrase superfamily (CA, EC 4.2.1.1) of metalloenzymes is present in all three domains of life (Eubacteria, Archaea, and Eukarya), being an interesting example of convergent/divergent evolution, with its seven families (α-, β-, γ-, δ-, ζ-, η-, and θ-CAs) described so far. CAs catalyse the simple, but physiologically crucial reaction of carbon dioxide hydration to bicarbonate and protons. Recently, our groups characterised the α-CA from the thermophilic bacterium, Sulfurihydrogenibium yellowstonense finding a very high catalytic activity for the CO2 hydration reaction (kcat = 9.35 × 105 s−1 and kcat/Km = 1.1 × 108 M−1 s−1) which was maintained after heating the enzyme at 80 °C for 3 h. This highly thermostable SspCA was covalently immobilised within polyurethane foam and onto the surface of magnetic Fe3O4 nanoparticles. Here, we describe a one-step procedure for immobilising the thermostable SspCA directly on the surface membrane of Escherichia coli, using the INPN domain of Pseudomonas syringae. This strategy has clear advantages with respect to other methods, which require as the first step the production and the purification of the biocatalyst, and as the second step the immobilisation of the enzyme onto a specific support. Our results demonstrate that thermostable SspCA fused to the INPN domain of P. syringae ice nucleation protein (INP) was correctly expressed on the outer membrane of engineered E. coli cells, affording for an easy approach to design biotechnological applications for this highly effective thermostable catalyst.

Cryoprotectant systems and cold tolerance of insects inhabiting central Yakutia (Russian Far East)

Strong tolerance of freezing is an important strategy for insects living in extremely cold regions. They produce highly effective cryoprotectant systems consisting of ice-nucleating proteins and polyols, which enables tolerable freezing of the body fluid. Therefore, the measurement of the concentrations of polyols and the activity of ice nucleators in the haemolymph is an essential tool for describing tolerance to ice formation in insects occurring in particularly cold places. This study evaluates three parameters: insect body supercooling point (SCP), haemolymph glycerol content and the profile of haemolymph ice nucleating activity that characterize the strategies of cold adaptation and cold hardiness in two previously unstudied beetles, Chrysolina graminis graminis L. and Galerucella nymphaea L., inhabiting Yakutia (Russian Far East, latitude 62°N). The high SCP values, ice nucleating activity and survival of the chrysomelids after freezing indicate that both species are tolerant of freezing. According to the profiles of ice-nucleating activity, the haemolymph from C. graminis graminis is characterized by a higher nucleating potential than that from G. nymphaea. The glycerol level is also higher in C. graminis graminis. The results indicate that both species develop tolerance to low temperatures, but the cold hardiness potential of C. graminis graminis is greater than that of G. nymphaea. This was revealed by the survival test, in which beetles were frozen to a temperature of -22°C for 30 min; 86% of C. graminis graminis and 72% of G. nymphaea survived the test. Thus, the freeze-tolerance of these beetles seems to be based on the production of an integrated cryoprotectant system, the quality of which apparently influences the range of their cold resistance.

Biophysical characterization of soluble Pseudomonas syringae ice nucleation protein InaZ fragments

Ice nucleation protein (INP) with its functional domain consisting of multiple 48-residue repeat units effectively induces super-cooled water into ice. Circular dichroism and infrared deconvolution analyses on a soluble 240-residue fragment of Pseudomonas syringae InaZ (InaZ240) containing five 48-residue repeat units indicated that it is mostly composed of β-sheet and random coil. Analytical ultracentrifugation suggested that InaZ240 behaves as a monomer of an elongated ellipsoid. However, InaZ240 showed only minimum ice binding compared to anti-freeze proteins. Other P. syringae InaZ proteins with more 48-residue repeat units were made, in which the largest soluble fragment obtainable was an InaZ with twelve 48-residue repeat units. Size-exclusion chromatography analyses further suggested that the overall shape of the expressed InaZ fragments is pH-dependent, which becomes compact as the numbers of 48-residue repeat unit increase.

Engineering Escherichia coli to bind to cyanobacteria

We engineered Escherichia coli cells to bind to cyanobacteria by heterologously producing and displaying lectins of the target cyanobacteria on their surface. To prove the efficacy of our approach, we tested this design on Microcystis aeruginosa with microvirin (Mvn), the lectin endogenously produced by this cyanobacterium. The coding sequence of Mvn was C-terminally fused to the ice nucleation protein NC (INPNC) gene and expressed in E.coli. Results showed that E.coli cells expressing the INPNC::Mvn fusion protein were able to bind to M.aeruginosa and the average number of E.coli cells bound to each cyanobacterial cell was enhanced 8-fold. Finally, a computational model was developed to simulate the binding reaction and help reconstruct the binding parameters. To our best knowledge, this is the first report on the binding of two organisms in liquid culture mediated by the surface display of lectins and it may serve as a novel approach to mediate microbial adhesion.