Review Macromolecular crowding: Macromolecules friend or foe Shruti Mittal, Rimpy Kaur Chowhan, Laishram Rajendrakumar Singh ⁎ Dr. B. R. Ambedkar Centre for Biomedical Research, University of Delhi, Delhi - 110 007, India abstract article info Article history: Received 13 March 2015 Received in revised form 27 April 2015 Accepted 4 May 2015 Available online 8 May 2015 Keywords: Protein folding Protein aggregation Enzyme activity Protein stability Excluded volume effect Background: Cellular interior is known to be densely crowded due to the presence of soluble and insoluble mac- romolecules, which altogether occupy ~40% of the total cellular volume. This results in altered biological proper- ties of macromolecules. Scope of Review: Macromolecular crowding is observed to have both positive and negative effects on protein fold- ing, structure, stability and function. Significant data has been accumulated so far on both the aspects. However, most of the review articles so far have focused on the positive aspect of macromolecular crowding and not much attention has been paid on the deleterious aspect of crowding on macromolecules. In order to have a complete knowledge of the effect of macromolecular crowding on proteins and enzymes, it is important to look into both the aspects of crowding to determine its precise role under physiological conditions. To fill the gap in the understanding of the effect of macromolecular crowding on proteins and enzymes, this review article focuses on the deleterious influence of crowding on macromolecules. Major Conclusions: Macromolecular crowding is not always good but also has several deleterious effects on var- ious macromolecular properties. Taken together, the properties of biological macromolecules in vivo appears to be finely regulated by the nature and level of the intracellular crowdedness in order to perform their biological functions appropriately. General Significance: The information provided here gives an understanding of the role played by the nature and level of cellular crowdedness in intensifying and/or alleviating the burden of various proteopathies. © 2015 Elsevier B.V. All rights reserved. 1. Introduction The present understanding of various biological processes has been acquired through investigations largely made under dilute experimen- tal conditions where the total macromolecular concentration never ex- ceeds 10 g/l. However, biological macromolecules are known to evolve and function under crowded intracellular environments consisting of a plethora of both soluble and insoluble macromolecules like proteins, nucleic acids, ribosomes and carbohydrates with their sum concentra- tion reaching around several hundred g/l. For example, the total concen- tration of protein and RNA inside the bacterium, Escherichia coli is in the range of 300–400 g/l [1]. Altogether, these macromolecules occupy a significant fraction (~40%) of the total cellular volume [2], making it vir- tually unavailable to the other macromolecules present. Such media are termed ‘crowded’ or ‘volume-occupied’ rather than ‘concentrated’, be- cause no single species of macromolecule is necessarily present at a high concentration. In fact, the level of crowdedness varies among different cell types and cellular compartments. Human lens contains ap- proximately 340 g/l protein [3]; the red blood cells contain about 350 g/l hemoglobin [4]; while the total protein content in the mitochondrial matrix may reach up to 500 g/l [5]. Macromolecular crowding is ob- served not only in the cellular interior but also in the extracellular ma- trix of tissues. For example, blood plasma contains ~80 g/l protein, a concentration high enough to cause significant crowding effects [2]. The degree of volume occupancy by these macromolecules is expected to have major thermodynamic and kinetic consequences on the proper- ties of macromolecules present in the cell [6–9]. The term ‘macromolec- ular crowding’ connotes the non-specific influence of steric repulsions on specific reactions, and processes that occur in highly volume- occupied media [10]. It was Minton and Wilf [11] who brought the influ- ence of crowding on macromolecules to the forefront in terms of theory and experiment and coined the term “macromolecular crowding” in 1981. The influence of macromolecular crowding on various properties of macromolecules has been examined in depth by adding high concentra- tions of inert synthetic or natural macromolecules, termed crowding agents or crowders, to the system in vitro to create an in vivo like sce- nario [2]. It is generally believed that macromolecular crowding (i) enhances protein stabilization against denaturation by heat, cold or denaturant. It has been argued that macromolecular crowding stabilizes globular proteins due to excluded volume effect because the native state occupies less space than the denatured state [12–15]; (ii) alters the re- action rates depending on the nature of reactions (diffusion-limited or transition-state-limited). Since, macromolecular crowding decreases Biochimica et Biophysica Acta 1850 (2015) 1822–1831 ⁎ Corresponding author. Tel.: +91 9811630757 (mobile). E-mail addresses: shrutimittal4@gmail.com (S. Mittal), rimpy_1989@yahoo.co.in (R.K. Chowhan), lrsingh@acbr.du.ac.in (L.R. Singh). http://dx.doi.org/10.1016/j.bbagen.2015.05.002 0304-4165/© 2015 Elsevier B.V. All rights reserved. Contents lists available at ScienceDirect Biochimica et Biophysica Acta journal homepage: www.elsevier.com/locate/bbagen