Chapter 8 1 Molecular Chaperones: Structure-Function 2 Relationship and their Role in Protein Folding 3 Bhaskar K. Chatterjee, Sarita Puri, Ashima Sharma, Ashutosh Pastor, 4 and Tapan K. Chaudhuri 5 Abstract During heat shock conditions a plethora of proteins are found to play a 6 role in maintaining cellular homeostasis. They play diverse roles from folding of 7 non-native proteins to the proteasomal degradation of harmful aggregates. A few 8 out of these heat shock proteins (Hsp) help in the folding of non-native substrate 9 proteins and are termed as molecular chaperones. Various structural and functional 10 adaptations make them work efficiently under both normal and stress conditions. 11 These adaptations involve transitions to oligomeric structures, thermal stability, 12 efficient binding affinity for substrates and co-chaperones, elevated synthesis during 13 shock conditions, switching between ‘holding’ and ‘folding’ functions etc. Their 14 ability to function under various kinds of stress conditions like heat shock, cancers, 15 neurodegenerative diseases, and in burdened cells due to recombinant protein pro- 16 duction makes them therapeutically and industrially important biomolecules. 17 Keywords Chaperone assisted folding · Heat shock · Molecular chaperones · 18 Protein folding · Structure-function of chaperones 19 Abbreviations 20 ACD α-crystallin domain 21 ADP Adenosine di-phosphate 22 ATP Adenosine tri-phosphate 23 CCT Chaperonin containing TCP-1 24 CIRCE Controlling inverted repeat of chaperone expression 25 Bhaskar K. Chatterjee, Sarita Puri, Ashima Sharma, and Ashutosh Pastor authors are equally contributed. B. K. Chatterjee · S. Puri · A. Sharma · A. Pastor · T. K. Chaudhuri (*) Kusuma School of Biological Sciences, Indian Institute of Technology Delhi, HauzKhas, New Delhi, India e-mail: [email protected] © Springer International Publishing AG 2018 181 A. A. A. Asea, P. Kaur (eds.), Regulation of Heat Shock Protein Responses, Heat Shock Proteins 14, https://doi.org/10.1007/978-3-319-74715-6_8 182 B. K. Chatterjee et al. 26 CNX Calnexin 27 CRT Calreticulin 28 CS Citrate synthase 29 ER Endoplasmic reticulum 30 ERAD Endoplasmic reticulum associated degradation 31 FRET Fluorescence energy resonance transfer 32 HOP HSP90/HSP70 organizing protein 33 HSC70 Heat shock cognate 34 HSEs Heat shock elements 35 HSFs Heat shock response and specific transcription factors 36 Hsp Heat shock proteins 37 HSP Heat shock protein family 38 HSR Heat shock response 39 MalZ Maltodextrin glucosidase 40 NAC Nascent chain associated complex 41 NEF Nucleotide-exchange factors 42 NTD n-terminal domain 43 PBD Peptide binding domain 44 PPIase Peptidyl-prolyl isomerases 45 PTP Permeability transition pore complex 46 RAC Ribosome associated complex 47 RuBisCO Ribulose-1,5-bisphosphate oxygenase-carboxylase 48 SHR Steroid hormone receptors 49 sHsp Small heat shock proteins 50 sHSP Small heat shock protein family 51 TF Trigger factor 52 TPR Tetratricopeptide 53 TRiC TCP-1 ring complex 54 UPR Unfolded protein response pathway 55 8.1 Introduction 56 Living systems respond to threatening conditions at multiple levels in their quest for 57 survival. It may in the form of a fight or flight response, which is a result of any 58 imminent physical threat either to an organism or their inner homeostasis. For 59 example the temperature, ionic and sugar balance are regulated within a fixed range 60 in our bodies and are probably optimized by evolutionary mechanisms. Similarly, 61 homeostasis is also maintained at the cellular level and maintaining such a balance 62 is imperative for the survival and efficient functioning of the cell. One of the major 63 homeostasis mechanisms operating at the cellular level is the protein homeostasis, 64 commonly referred to as proteostasis (Balch et al. 2008). Starting with maintaining 65 the structural organization of a cell to catalysing various metabolic reactions; from 66 the transport of macromolecules within and across cells to various recognition and 8 Molecular Chaperones in Cellular Stress Response 183 immune functions, proteins play vital roles in our bodies and are regarded as the 67 actual workhorses of the cells. Proteins undergo various post-translational modifica- 68 tions and move through trafficking pathways before they are ready to take up their 69 function. However, acquiring their specific three-dimensional structure supersedes 70 all this because only in their specific structural forms can they undertake the func- 71 tion they are meant to carry out. In this chapter, we shall discuss the cellular machin- 72 ery responsible for maintaining proteins in their functional state during stress 73 conditions; specifically focusing on the Hsp that assist in the folding and refolding 74 of misfolded and aggregation prone proteins. 75 8.2 What Is Stress Response? 76 The stress response can be defined as our involuntary defense reaction to threaten- 77 ing conditions. This may occur at multiple levels as a response to a different range 78 of conditions. At a cellular level, the response to any alarming condition, like a 79 chronic change in the environment away from normal conditions (which may inter- 80 fere with the physiological functioning of the cells and cause damage to nucleic 81 acids and proteins) can be identified as a stress response (Kültz 2004). While at the 82 organism level our adaptive responses are driven by hormonal changes (Charmandari 83 et al. 2005), the cellular response to damages occurring at a molecular level involves 84 a cascade of pathways, and molecules that work in cohort to bring the cells back to 85 their normal functional state. Various kinds of stress at the cellular level include 86 oxidative, heat, radiation and nutrient deprivation. The major consequences of these 87 stress are DNA damage, loss of cellular signalling, protein unfolding, misfolding, 88 aggregation, proteolysis, cellular necrosis and apoptosis. The cellular stress response 89 may be of a protective nature where the cell can defend and restore its normal func- 90 tioning, or of a destructive nature where the conditions are beyond the cell’s ability 91 to repair. The type, level, and duration of stressful conditions may ultimately deter- 92 mine the fate a cell. During stress conditions, proteins are misfolded due to changes 93 in the overall energy landscape. This causes loss of protein function and the accu- 94 mulation of misfolded proteins in the form of toxic aggregates. The protective stress 95 response for proteins includes pathways of the heat shock response and the unfolded 96 protein response (UPR); the destructive response pathways include apoptosis, 97 necrosis or autophagy (Fulda et al. 2010). The DNA damage response consists of 98 multiple, complex pathways that restore genomic integrity. These include the base 99 excision repair, nucleotide excision repair, and non-homologous end joining 100 (Kourtis and Tavernarakis 2011). The oxidative stress response helps the cell cope 101 with the reactive oxygen species, maintain redox homeostasis; and a number of 102 enzymes like superoxide dismutase and non-enzymatic antioxidants are involved 103 (Trachootham et al. 2008). In this chapter, we shall mainly focus on the diverse 104 mechanisms governing heat shock response and the various factors that are involved 105 in mediating such a response. 106 184 B. K. Chatterjee et al. 107 8.3 Heat Shock Response 108 The heat shock response was one of the earliest explored stress response mecha- 109 nisms that was initially observed in Drosophila as changes in puffing patterns of 110 salivary gland chromosomes (Ritossa 1996), followed by changes in gene expres- 111 sion patterns after heat treatment (Tissiéres et al. 1974; Hightower 1991). The heat 112 shock response imparts thermo-tolerance to the cells and protects them when they 113 are stressed due to prolonged exposure to heat. This response is activated by the rise 114 of a few degrees in temperature from the normal dwelling temperature of the organ- 115 ism. The regular transcription and translation processes in the cells are halted during 116 heat shock response, and specific transcription factors (HSFs) which selectively 117 enhance expression of a set of proteins having protective functions are activated. 118 However, even during normal conditions these HSFs play important roles in differ- 119 entiation and development of the organisms (Morimoto et al. 1996). The HSF1 120 predominantly regulates the heat shock responses, and is itself regulated by its 121 interactions with heat shock proteins HSP70 and HSP90 (Pirkkala et al. 2001). The 122 ability to activate transcription and bind to DNA are uncoupled in HSF1 imparting 123 a higher degree of regulation. The HSF1 exists as a monomer or in complex with 124 Hsp under normal conditions. During heat shock, HSF1 homotrimerizes and under- 125 goes hyperphosphorylation which leads to its activation (Cotto et al. 1996). These 126 HSFs facilitate the overexpression of Hsp by binding to cis acting sequences on the 127 genome known as heat shock elements (HSEs). HSP are commonly known as 128 molecular chaperones for their role in assisting proteins to acquire their native struc- 129 tures. Hsp prevent heat induced denaturation and aggregation of proteins, facilitate 130 the proteins to fold, and assist in the refolding of already denatured proteins 131 (Lindquist 1986). The Hsp play an active role in facilitating the degradation of 132 proteins that are unable to fold in order to maintain protein homeostasis and thus 133 promote cell survival. 134 8.4 Cellular Components Providing HS Response 135 Heat shock response in cells is mediated by concerted actions of heat shock factors 136 (HSF), heat shock elements (HSE) and Hsp. The heat shock factors as described 137 above are transcription factors activated during a heat shock. Gram negative bacte- 138 ria E.coli has a specific sigma factor 32 σ( 32), coded by the rpoH gene, which is a 139 heat shock promoter specific subunit of RNA polymerase. Theσ 32 is a positive 140 regulator and is suppressed by DnaJ during normal conditions (Bukau 1993).