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Itaru Urabe

Publications and source records attributed to Itaru Urabe.

5 recordsLinked to original sources
PubMed2003

Can an arbitrary sequence evolve towards acquiring a biological function?

To explore the possibility that an arbitrary sequence can evolve towards acquiring functional role when fused with other pre-existing protein modules, we replaced the D2 domain of the fd-tet phage genome with the soluble random polypeptide RP3-42. The replacement yielded an fd-RP defective phage that is six-order magnitude lower infectivity than the wild-type fd-tet phage. The evolvability of RP3-42 was investigated through iterative mutation and selection. Each generation consists of a maximum of ten arbitrarily chosen clones, whereby the clone with highest infectivity was selected to be the parent clone of the generation that followed. The experimental evolution attested that, from an initial single random sequence, there will be selectable variation in a property of interest and that the property in question was able to improve over several generations. fd-7, the clone with highest infectivity at the end of the experimental evolution, showed a 240-fold increase in infectivity as compared to its origin, fd-RP. Analysis by phage ELISA using anti-M13 antibody and anti-T7 antibody revealed that about 37-fold increase in the infectivity of fd-7 was attributed to the changes in the molecular property of the single polypeptide that replaced the D2 domain of the g3p protein. This study therefore exemplifies the process of a random polypeptide generating a functional role in rejuvenating the infectivity of a defective bacteriophage when fused to some preexisting protein modules, indicating that an arbitrary sequence can evolve toward acquiring a functional role. Overall, this study could herald the conception of new perspective regarding primordial polypeptides in the field of molecular evolution.

Amino Acid Sequence↗
PubMed2002

Importance of compartment formation for a self-encoding system.

A self-encoding system designed to have strict "compartition" of the molecules, i.e., to contain only a single molecule of DNA in each compartment, was established, and its evolutionary fate was analyzed. The system comprised the Thermus thermophilus DNA polymerase gene as the informational molecule and its protein product replicating the gene as the functional molecule. Imposing strict compartition allows the self-encoding system to last up to at least the tenth generation, whereas the system ceased to work after the third generation when loose compartition initiated with 100 molecules was imposed. These results provide experimental evidence on the importance of compartition for the maintenance of a self-encoding system. In addition, the extent of diversity in self-replication activity of the compartments was found to be another vital difference in the evolutionary dynamics between the strict and loose compartitions. Although the system with strict compartition provides widely diversified activity of the compartments at each generation, the values of the activity diverge only within a small range in the system with loose compartition. When the variety in the activity of a compartment is small, functional selection becomes weak, and to conform Darwinian evolution may become unfeasible. Therefore, strict compartition is essential for the evolvability of a self-encoding system.

Amino Acid Substitution↗
PubMed2002

Evolvability of random polypeptides through functional selection within a small library.

A directed evolution with phage-displayed random polypeptides of about 140 amino acid residues was followed until the sixth generation under a selection based on affinity to a transition state analog for an esterase reaction. The experimental design deliberately limits the observation to only 10 clones per generation. The first generation consists of three soluble random polypeptides and seven arbitrarily chosen clones from a previously constructed library. The clone showing the highest affinity in a generation was selected and subjected to random mutagenesis to generate variants for the next generation. Even within only 10 arbitrarily chosen polypeptides in each of the generations, there are enough variants in accord to capacity of binding affinity. In addition, the binding capacity of the selected polypeptides showed a gradual continuous increase over the generation. Furthermore, the purified selected random polypeptides exhibited a gradual but significant increase in esterase activity. The ease of the functional development within a small sequence variety implies that enzyme evolution is prompted even within a small population of random polypeptides.

Amino Acid Sequence↗
PubMedDate not supplied

An observation of the initial stage towards a symbiotic relationship.

Two well-characterized and phylogeneticaly different species, Escherichia coli and Dictyostelium discoideum, were used as the model organisms. When the two species were mixed and allowed to grow on minimal agar plates at 22 degrees C, remarkably, the two species achieved a state of coexistence at an average of 2-4 weeks. In addition, the emerged colonies housing the coexisting species had a mucoidal nature that was not observed from its origin. Moreover, the state of coexistence was confirmed to be stable, and so was the mucoidal nature of the emerged colonies. Comparing with the pure E. coli origin, the mucoidal colony showed a significant increase in higher molecular weight extracellular components, with polysaccharides as the major constituent. Qualitative analysis of the monosaccharide contents in the extracellular components of the mucoidal colony revealed not only a significant increase in the glucose content, but also significant amount of additional xylose and galactose. The system permits the initial stages of the development of relationship between two species be captured within a short period of time. This feature, together with being simple and reproducible in laboratory conditions, provides a new model system for the study of symbiosis, especially when initial stages are concerned.

Animals↗