Pier Luigi Luisi

• Roma Tre University

It is well known that life is cellular and only cellular: all tissues and organs of all animals and plants are organized assemblies of cells—so that we can consider the cell as the elemental constituent of life on this planet.

Primitive cell models help to understand the role that compartmentalization plays in origin of life scenarios. Here we present a combined experimental and modeling approach towards the construction of simple model systems for primitive cellular assemblies. Charged lipid vesicles aggregate in the presence of oppositely charged biopolymers, such as n...

The cover picture shows a confocal fluorescence picture of water-in-oil microemulsion droplets synthesizing enhanced green fluorescent protein by cell-free E. coli extracts-an in vitro compartmentation (IVC) system. As shown by the dynamic light-scattering profile, most of the droplets have radius of around 300-500 nm. The droplet interface is comp...

The spontaneous formation of lipid vesicles, in particular fatty acid vesicles, is considered an important physical process at the roots of cellular life. It has been demonstrated previously that the addition of fatty acid micelles to preformed vesicles induces vesicle self-reproduction by a growth-division mechanism. Despite multiple experimental...

Water-in-oil (w/o) emulsions are used as a cellular model because of their unique cell-like architecture. Previous works showed the capability of eukaryotic-cell-sized w/o droplets (5–50 μm) to support protein synthesis efficiently; however data about smaller w/o compartments (<1 μm) are lacking. This work focuses on the biosynthesis of the enhance...

This analysis starts from the view that prebiotic chemical evolution, leading to the first forms of life on earth, was based on a series of sequential steps, each determined by its own contingent initial conditions. This view is opposed to the more established one, which sees the origin of life as a series of preordered series of events, where each...

How did primitive living cells originate? The formation of early cells, which were probably solute-filled vesicles capable of performing a rudimentary metabolism (and possibly self-reproduction), is still one of the big unsolved questions in origin of life. We have recently used lipid vesicles (liposomes) as primitive cell models, aiming at the stu...

It is pointed out that one of the main reasons of lack of real conceptual progress in the field may lie in the fact that questions concerning the biogenesis of macromolecules have never been asked or addressed in a proper way. We should start again research on the origin of life starting from "ground zero" and focusing on the prebiotic synthesis of...

We emphasize here that, in considering the initial prebiotic reactions, it is fundamental to take into consideration the critical threshold concentration, in particular when talking about self-replication and initial metabolism. It is also shown that the in situ formation of vesicles in a solution containing macromolecular solutes, permits to obtai...

The question about the origin of life is still largely unanswered, is in fact one of the great mysteries that science is still facing. We all accept the 1924 idea of Oparin, according to which life originated from the inanimate matter through a long series of steps of increasing molecular complexity and functionality. And we accept also that the fi...

It is argued that closed, cell-like compartments, may have existed in prebiotic time, showing a simplified metabolism which was bringing about a primitive form of stationary state- a kind of homeostasis. The autopoietic primitive cell can be taken as an example and there are preliminary experimental data supporting the possible existence of this pr...

One of the main open questions in origin of life research focuses on the formation, by self-organization, of primitive cells composed by macromolecular compounds enclosed within a semi-permeable membrane. A successful experimental strategy for studying the emergence and the properties of primitive cells relies on a synthetic biology approach, consi...

Here we report some recent biophysical issues on the preparation of solute-filled lipid vesicles and their relevance to the construction of “synthetic cells.” First, we introduce the “semi-synthetic minimal cells” as the liposome-based cell-like systems, which contain a minimal number of biomolecules required to display simple and complex biologica...

A prebiotic origin of metabolism has been proposed as one of several scenarios for the origin of life. In their recent work, Ralser and colleagues (Keller et al, ) observe an enzyme-free, metabolism-like reaction network under conditions reproducing a possible prebiotic environment. A prebiotic origin of metabolism has been proposed as one of sever...

Although both the most popular form of synthetic biology (SB) and chemical synthetic biology (CSB) share the biotechnologically useful aim of making new forms of life, SB does so by using genetic manipulation of extant microorganism, while CSB utilises classic chemical procedures in order to obtain biological structures which are non-existent in na...

In this mini-review we present some experimental approaches to the important issue in the origin of life, namely the origin of nucleic acids and proteins with specific and functional sequences. The formation of macromolecules on prebiotic Earth faces practical and conceptual difficulties. From the chemical viewpoint, macromolecules are formed by ch...

The recent advancements in semi-synthetic minimal cell (SSMC) technology pave the way for several interesting scenarios that span from basic scientific advancements to applications in biotechnology. In this short chapter we discuss the relevance of establishing chemical communication between synthetic and natural cells as an important conceptual is...

Encapsulation: The emergence of primitive cells remains an enigma of the origin of life. By modeling this key process as the encapsulation of a complex multimolecular mixture inside liposomes, a remarkable self-organization process has been revealed that brings about solute-rich compartments in which protein synthesis can take place.

This is a report of a teaching course in a Buddhist monastery (a nunnery) in Bhutan, where I—as a member of the Midn and Life Institute directed by the Dalai Lama—have been invited to illustrate the main aspects of modern science in the field of the origin of life and evolution. The class was formed by ca. 60 nuns, and a good number of Buddhist Lam...

Chemical synthetic biology (CSB) is a branch of synthetic biology (SB) oriented toward the synthesis of chemical structures alternative to those present in nature. Whereas SB combines biology and engineering with the aim of synthesizing biological structures or life forms that do not exist in nature – often based on genome manipulation, CSB uses an...

Background The wet-lab synthesis of the simplest forms of life (minimal cells) is a challenging aspect in modern synthetic biology. Quasi-cellular systems able to produce proteins directly from DNA can be obtained by encapsulating the cell-free transcription/translation system PURESYSTEM™(PS) in liposomes. It is possible to detect the intra-vesicle...

The notion of minimal cells refers to cellular structures that contain the minimal and sufficient complexity to still be defined as living, or at least capable to display the most important features of biological cells. Here we briefly describe the laboratory construction of minimal cells, a project within the broader field of synthetic biology. In...

A new scenario for prebiotic formation of nucleic acid oligomers is presented. Peptide catalysis is applied to achieve condensation of activated RNA monomers into short RNA chains. As catalysts, L-dipeptides containing a histidine residue, primarily Ser-His, were used. Reactions were carried out in selforganised environment, a water-ice eutectic ph...

The inside cover picture shows the oligomerization of activated RNA monomers by the catalytic dipeptide Ser‐His. On p. 217 ff., P.‐A. Monnard et al. explain how the reaction takes place in a water/ice eutectic phase below 0 °C. This reaction environment is believed to be prebiotically plausible and likely present today on various planetary bodies l...

The construction of synthetic cells is one of the major goals of bioengineering. The most successful approach consists in the encapsulation of biochemical materials (DNA, RNA, enzymes, etc.) inside lipid vesicles (liposomes), mimicking a cell structure. In this contribution, that also aims at introducing the reader to 'chemical synthetic biology,'...

The general framework of the origin of life on Earth is outlined, emphasizing that the so-called prebiotic 'RNA world' is as yet on shaky scientific ground, and that one should any way ask the question of the structure of the first protocellular compartments capable of the initial forms of metabolism. This question is the basis of the research proj...

Here we summarize the main results of our latest investigation on the spontaneous encapsulation of proteins (ferritin) and ribosomes inside lipid vesicles. We show that when vesicles form in a solution containing some macromolecules (even at low concentration), in contrast to the expectations, a few but measurable number of vesicles is able to capt...

Current research on the origin of life typically focuses on the self-organisation of molecular components in individual cell-like compartments, thereby bringing about the emergence of self-sustaining minimal cells. This is justified by the fact that single cells are the minimal forms of life. No attempts have been made to investigate the cooperativ...

Retinoids represent the first-line therapy for the treatment of acne vulgaris. Their effect is comedolytic and anti-comedogenic, and associates with hyperplasia and deregulated differentiation of the epidermis, and decreased inflammation. We here tested the comedolytic effect of the novel atypical retinoid E-3-(3'-Adamantan-1-yl-4'-methoxybiphenyl-...

Synthetic biology is first represented in terms of two complementary aspects, the bio-engineering one, based on the genetic manipulation of extant microbial forms in order to obtain forms of life which do not exist in nature; and the chemical synthetic biology, an approach mostly based on chemical manipulation for the laboratory synthesis of biolog...

Biological systems evolved with the ability to communicate with their biotic surroundings through chemical signalling. Production, perception and decoding of the information carried by signal molecules allow individuals of a community to interact, cooperate, and coordinate their activities, establishing complex social behaviours. In this paper we s...

Over the past thirty years, a new systemic conception of life has emerged at the forefront of science. New emphasis has been given to complexity, networks, and patterns of organisation leading to a novel kind of 'systemic' thinking. This volume integrates the ideas, models, and theories underlying the systems view of life into a single coherent fra...

In this mini-review we would like to summarize the recent advances in the field of protein and nucleic acid synthesis inside lipidvesicles (liposomes). This research, which originated within the origin of life community, is now recognized as an example of synthetic biology. Current approaches are based on the convergence of liposome technology and...

I review here my personal and scientific interactions with Francisco Varela, starting from our meeting in 1983 in Alpbach, Austria, a momentous meeting, which was also the place where the Mind and Life Institute and independently the Cortona week were conceived. Later on, the scientific cooperation focussed on autopoiesis and permitted to arrive at...

One of the open questions in the origin of life is the spontaneous formation of primitive cell-like compartments from free molecules in solution and membranes. "Metabolism-first" and "replicator-first" theories claim that early catalytic cycles first evolved in solution, and became encapsulated inside lipid vesicles later on. "Compartment-first" th...

In this chapter we present our “semi-synthetic” approach to the construction of minimal living cells. Firstly, we shortly review the advancements carried out in the last few years, especially in the field of reactions inside liposomes. Then we discuss our recent study on the investigation of minimal cell size, carried out from the perspective of sy...

The self-reproduction of a giant lipid vesicle has been linked to the replication of encapsulated DNA -- a promising combination for the construction of a minimalistic synthetic cell.

In the last years, we have developed a method for the study of random RNA sequences with the aim of investigating their capacity to assume folded and possibly functionally active (ribozymes) structures. The RNA Foster assay developed in our laboratory is a powerful, simple, and fast method to investigate the structural properties of RNA by assessin...

Compartmentalization is a necessary element for the development of any cell cycle and the origin of speciation. Changes in shape and size of compartments might have been the first manifestation of development of so-called cell cycles. Cell growth and division, processes guided by biological reactions in modern cells, might have originated as purely...

In this chapter we describe the concept of minimal living cells, defined as synthetic or semi-synthetic cells having the minimal and sufficient number of components to be endowed with the main biological properties of living cells. The construction of minimal cells starting from isolated compounds is an issue in synthetic biology, origins of life s...

This chapter is an attempt to provide a framework of epistemology to the growing field of synthetic biology (SB). It is preliminarily argued that there are two kinds of SB, one (bioengineering SB) clearly and purposely directed towards one goal set from the start and the other kind being more concerned with basic science and responding to the basic...

The Notion of Minimal CellThe Minimal GenomeThe Minimal RNA CellThe Minimal Size of CellsCurrent Experimental Approaches for the Construction of Minimal CellsConcluding RemarksAcknowledgmentsReference

IntroductionRandom RNAs Readily Adopt a Compact and Thermostable Secondary StructureRandom RNAs can be Easily Designed and Engineered to Acquire Novel FunctionsRandom RNAs as a Modular Scaffold for Synthetic BiologyConclusions AcknowledgmentsReferences

Chemistry plays a very important role in the emerging field of synthetic biology. In particular, chemical synthetic biology is concerned with the synthesis of chemical structures, such as proteins, that do not exist in nature. With contributions from leading international experts, Chemical Synthetic Biology shows how chemistry underpins synthetic b...

Three examples of this "chemical synthetic biology" approach are given in this article. The first example deals with the synthesis of proteins that do not exist in nature, and dubbed as "the never born proteins" (NBPs). This research is related to the question why and how the protein structures existing in our world have been selected out, with the...

In the last ten years there has been a considerable increase of interest on the notion of the minimal cell. With this term we usually mean a cell-like structure containing the minimal and sufficient number of components to be defined as alive, or at least capable of displaying some of the fundamental functions of a living cell. In fact, when we loo...

ChemInform is a weekly Abstracting Service, delivering concise information at a glance that was extracted from about 100 leading journals. To access a ChemInform Abstract of an article which was published elsewhere, please select a “Full Text” option. The original article is trackable via the “References” option.

Micelles and vesicles display a few characteristic features which are present in the structures of the living. First of all, they are self-assembling, i.e. they are examples of self-organization which is under thermodynamic control. Furthermore, micelles and vesicles are bounded structures, i.e. they have a closed interface (boundary) which discrim...

Bringing together the latest scientific advances and some of the most enduring subtle philosophical puzzles and problems, this book collects original historical and contemporary sources to explore the wide range of issues surrounding the nature of life. Selections ranging from Aristotle and Descartes to Sagan and Dawkins are organised around four b...

(Figure Presented) Ferritin encapsulation inside lipid vesicles reveals the spontaneous formation of protein-rich vesicles. The solute distribution inside the vesicles follows a power law. The important conclusion for origins-of-life scenarios is that the dynamics of membrane closure allow the accumulation of solutes inside primitive cells, thus pr...

ChemInform is a weekly Abstracting Service, delivering concise information at a glance that was extracted from about 100 leading journals. To access a ChemInform Abstract of an article which was published elsewhere, please select a “Full Text” option. The original article is trackable via the “References” option.

IntroductionLipid Vesicles (Liposomes)Experimental Strategies and Theoretical AspectsA Theoretical Framework for Complex Reactions in LiposomesFour Cases of Compartmentalized ReactionsConclusion AcknowledgementsAbbreviationsReferences

Supramolecular chemistry was enriched, about twenty years ago, by the discovery of the self-reproduction of micelles and vesicles. The dynamic aspects and complexity of these systems makes them good models for biological compartments. For example, the self-reproduction of vesicles suggests that the growth in size and number of a vesicle population...

In this paper we develop the autopoietic approach to the definition of the living developed by Maturana and Varela in the Seventies. Starting from very simple observations concerning the phenomenology of life, we propose a reformulation of the autopoietic original definition of life which integrates some of the contemporary criticism to it. Our def...

The recent advent, success and diffusion of synthetic biology (SB) are mainly related to its application as markedly bioengineering-oriented discipline. In addition to this classical view, SB also means "constructive" biology, and it is aimed to the construction of synthetic (artificial, man-made) biological-like systems, at the aim of understandin...

Protein expression is the most complex metabolic reaction that has been encapsulated in liposomes, mainly as an intermediate step toward the synthesis of minimal semisynthetic cells. Although there are different experimental approaches to achieving the synthesis of proteins inside liposomes and it is therefore not possible to give a standard recipe...

The term chemical synthetic biology defines that part of the field that, instead of assuming an engineering approach based on genome manipulation, is oriented towards the synthesis of chemical structures alternative to those present in nature. Several different literature projects will be illustrated, including the two of our group. One is concerne...

The hydrolytic activity of lipolytic enzymes in reverse micelles can be measured continuously with Fourier Transform infrared spectroscopy (FTIR) by following in the region of the OH-stretching band the water consumption during the reaction. This possibility is unique to reverse micellar solutions, because they are optically transparent and because...

Reverse micelles formed by soybean lecithin in isooctane were used as a reaction medium for both the lipase-catalyzed hydrolysis as well as the synthesis of lipids. Neither reaction appears to follow Michaelis-Menten kinetics and it is suggested that the rates are diffusion controlled. The hydrolysis of para-nitrophenylpalmitate (PNPP) and, in part...

This paper describes a novel environment for enzymatic reactions, a phospholipid-type matrix simulating a biological lipidic structure, which allows direct measurement of reaction parameters by noninvasive spectroscopic means.

The question of the minimal size of a cell that is still capable of endorsing life has been discussed extensively in the literature, but it has not been tackled experimentally by a synthetic-biology approach. This is the aim of the present work; in particular, we examined the question of the minimal physical size of cells using liposomes that entra...

The cover picture shows the concept of semisynthetic cell construction by co- entrapping DNA, enzymes, ribosomes, tRNAs, and small compounds (amino acids, NTPs, etc.) within a lipid vesicle. In order to be able to synthesize a protein, all components of the translation–transcription machinery must be present in the compartment, which is formed spon...

Synthetic biology is an emerging field that aims at constructing artificial biological systems by combining engineering and molecular biology approaches. One of the most ambitious research line concerns the so-called semi-synthetic minimal cells, which are liposome-based system capable of synthesizing the lipids within the liposome surface. This go...

The number of natural proteins represents a small fraction of all the possible protein sequences and there is an enormous number of proteins never sampled by nature, the so called "never born proteins" (NBPs). A fundamental question in this regard is if the ensemble of natural proteins possesses peculiar chemical and physical properties or if it is...

The dipeptide seryl-histidine (Ser-His) catalyses the condensation of esters of amino acids, peptide fragments, and peptide nucleic acid (PNA) building blocks, bringing to the formation of peptide bonds. Di-, tri- or tetra-peptides can be formed with yields that vary from 0.5% to 60% depending on the nature of the substrate and on the conditions. O...

The number of natural protein sequences is infinitely small as compared to the number of proteins theoretically possible. Thus, a huge number of protein sequences, defined as “never born proteins” or NBPs, have never been observed in nature. The study of the structural and functional properties of NBPs represents a way to improve our knowledge on t...

The discovery of self-reproduction of micelles and vesicles about 15 years ago opened a new page in the field of the supramolecular chemistry of surfactant aggregation phenomena, both for the importance of dynamic aspects of complexity of these systems and for its biological meaning. In fact, the self-reproduction of vesicles has suggested that the...

The first comprehensive general resource on state-of-the-art protocell research, describing current approaches to making new forms of life from scratch in the laboratory. Protocells offers a comprehensive resource on current attempts to create simple forms of life from scratch in the laboratory. These minimal versions of cells, known as protocells,...

Abstract As a continuation of our endeavor to find conditions under which bounded aggregate structures are able to self-reproduce, we have investigated the reactivity of lipase, both in free solution and vesicle-entrapped, against mixed oleic acid/oleate/ethyl oleate vesicles. Three types of vesicles have been prepared and characterized: (A) oleic...