The Pentameric Vertex Proteins Are Necessary for the Icosahedral Carboxysome Shell to Function as a CO2 Leakage Barrier

Abstract

BackgroundCarboxysomes are polyhedral protein microcompartments found in many autotrophic bacteria; they encapsulate the CO2 fixing enzyme, ribulose-1,5-bisphosphate carboxylase/oxygenase (RubisCO) within a thin protein shell and provide an environment that enhances the catalytic capabilities of the enzyme. Two types of shell protein constituents are common to carboxysomes and related microcompartments of heterotrophic bacteria, and the genes for these proteins are found in a large variety of bacteria.Methodology/Principal FindingsWe have created a Halothiobacillus neapolitanus knockout mutant that does not produce the two paralogous CsoS4 proteins thought to occupy the vertices of the icosahedral carboxysomes and related microcompartments. Biochemical and ultrastructural analyses indicated that the mutant predominantly forms carboxysomes of normal appearance, in addition to some elongated microcompartments. Despite their normal shape, purified mutant carboxysomes are functionally impaired, although the activities of the encapsulated enzymes are not negatively affected.Conclusions/SignificanceIn the absence of the CsoS4 proteins the carboxysome shell loses its limited permeability to CO2 and is no longer able to provide the catalytic advantage RubisCO derives from microcompartmentalization. This study presents direct evidence that the diffusion barrier property of the carboxysome shell contributes significantly to the biological function of the carboxysome.

Functions, Compositions, and Evolution of the Two Types of Carboxysomes: Polyhedral Microcompartments That Facilitate CO 2 Fixation in Cyanobacteria and Some Proteobacteria

“…Basic concepts inherent in early models of carboxysome organization and function (88)(89)(90) have largely been borne out by structurefunction analyses. Reinhold and coworkers' most developed model specifically placed the CO 2 diffusion resistance function at the carboxysome shell (88), and this function was subsequently confirmed in both ␣-and ␤-carboxysomes (19,(91)(92)(93)(94). Similarly, current models of carboxysomal CA localization, where CA nuclei are present throughout the shell structure (95)(96)(97)(98)(99), are consistent with one model version (88) in which numerous "CA sites" are surrounded by "RubisCO zones" rather than carboxysomes which contain a monolithic CA core.…”

Diverse Bacterial Microcompartment Organelles

“…Genetic studies in several systems showed that a known pentamer or a pentamer homolog is required for MCP formation in vivo (48,58,72). In another study, the pentameric protein, CsoS4, of the ␣-carboxysome was not absolutely required for shell assembly but was required for shell closure and integrity (73). In another case, a conflicting result was initially obtained with regard to a pentamer homolog encoded by the eut operon.…”

Bacterial Microcompartments

“…Coenzyme B 12 A complex organometallic cofactor associated with enzymes central to the degradation pathways of ethanolamine and 1,2-propandiol. …”