**1. Introduction**

Bioinformatics is an interdisciplinary science that uses both computational and informational approaches to retrieve, analyze, organize, visualize, store and develop biological data [1]. It is widely applied in the field of life sciences, especially in functional genomics, sequence analysis, proteomics, drug discovery, etc. Prediction of the structure and functions of the genes and proteins have become a fundamental task in the life science researches. The present book chapter involves molecular modeling study to investigate intermolecular interactions between Microtubule (MT) and Tau. Though these interactions are important in Alzheimer's disease, the

detailed knowledge on MT-Tau interactions are still lacking mainly because of two reasons (i) Lack of full-length structure of tau due to its intrinsically disordered nature and (ii) Differential expression of tubulin isotypes in different type of cells in particular brain and neuronal cells. Earlier experimental efforts have been made to elucidate these interactions using solution structure (PDB ID: 2MZ7.pdb) however correct binding mode and atomistic interactions at structural level are poorly understood. Therefore, this chapter focuses on application of molecular modeling techniques in understanding important MT-Tau interactions in the Alzheimers disease. Bioinformatics approaches like sequence analysis, homology modeling. MD simulations and binding energy calculations are employed systematically to address this challenging problem in the field of Alzheimer's disease.

Tau is intrinsically disordered protein encoded by '*mapt'* gene located on chromosome 17 [2]. The primary function of the tau protein is to bind and stabilize the microtubule. It is abundantly expressed in the brain and neuronal tissues hence its misregulation is associated with the Alzheimers and other neurodegenerative disorders [3, 4]. Till date about six isoforms of tau are reported in the human central nervous system. The length of these six isoforms varies between 352 to 441 residues [5].

Primary structure of tau contains the projectile domain at N-terminal (residue 1–244) which is composed of the acidic and proline-rich region, and the C-terminal repeat domain which consists of 4 repeats i.e., R1, R2, R3 and R4 (residues 245–441) (**Figure 1**). The six isoforms of tau mainly differs by the existence of either R3 or R4 repeats at the C-terminal domain [6]. The one of the tau isoforms is referred as longest isoform mostly observed in humans which comprises 4 repeats i.e., R1, R2, R3 and R4. while the shortest isoform of tau has only 3 repeats (R1, R2 and R3). This shortest isoform of tau is reported in the fetus brain and less common in adults [5, 7]. **Figure 1A** represents the structure of tau repeat region R2 which is bound to the MT composed of β/α/β tubulin subunits [8]. Hereafter, tau repeat R2 will be mentioned as 'TauR2' for the simplicity. The tau repeats R1, R2, R3 and R4 prefers to bind at the outer surface of microtubule (MT) to stabilize it (**Figure 1A**) and regulates MT polymerization [6]. **Figure 1B** represents domain organization in the tau primary structure and **Figure 1C** shows the sequence of TauR2 which is reported in the CryoEM model. It is well established that tau primarily helps in the assembly and stabilization of axonal MTs, which contributes to the proper functioning of neuronal cells [9]. However, recent studies have reported new functional role of the tau in addition to the axonal, i.e. labile domain of the MT to promote its assembly [10]. Tau detaches from the MTs and forms abnormal, fibrillar structures of insoluble aggregates due to post-translational modifications in Alzheimer diseases and other neurodegenerative diseases associated with tau [11, 12].

Full-length structure of tau protein is not yet determined using X-ray crystallographic techniques due to its intrinsically disordered nature. Also, the efforts to find its solution structure using NMR spectroscopy have failed [13]. Thus, the MT-Tau interactions have been studied so far using various biochemical and biophysical techniques [14–17]. The CryoEM have showed marginal success in determining the structure of tau repeat R2 bound to MT however it shows discontinues density of tau repeats along with each protofilament upon MT binding [8]. Hence, they synthetically developed R1 and R2 repeat of tau and their interactions with MT were examined. These two tau repeats adopts the extended conformation along the crest of protofilament which stabilizes the MT structure by binding to the interface of tubulin dimers [8].

MTs are made from αβ-tubulin heterodimer subunits [18]. In human, seven α-tubulin and nine β-tubulin isotypes are reported showing their tissue-specific expressions. For instance, βI tubulin isotype is ubiquitously expressed in all cells,

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**Figure 1.**

*Homology Modeling of Tubulin Isotypes to Investigate MT-Tau Interactions*

βII and βIII tubulin isotype are mainly expressed in brain and neuronal cells, and βVI tubulin isotype is expressed in erythroid cells and platelets [19]. The βI tubulin isotype reported to play crucial role in cell viability, βII tubulin isotype is important for neurite growth and βIII tubulin isotype protects nerve cell against free radicals and reactive oxygen species [20]. It has been well known that all β-tubulin isotypes share a significant residue conservation except the C-terminal tail region of MT [21–24] which is flexible in nature and structurally disordered. The C- tail region of all these isotypes overhang outwards of the MTs. The C-tail shows interactions with

*CryoEM Structure of tubulin subunits bound to TauR2. (A) tubulin subunits bound to TauR2 in CryoEM structure 6CVN.pdb. TauR2 domain binds at the outer surface of the MT. (B) Domain organization in tau,* 

It is well documented that the composition of β-tubulin isotypes (i) affects MT dynamic instability [27, 28], (ii) their interaction with motor proteins [29], (iii) their binding to the anti-drugs [21, 22, 30] and (iv) different MAPs including tau [31, 32]. These tubulin isotypes show tissue specific expression as their relative proportion varies greatly in different type of cells [20, 33, 34]. It is also well established that binding of tau to the MT promote or demote microtubule polymerization [35]. However, the differential binding affinity of tau to the various β-tubulin isotypes

various MAPs including tau and regulate MT dynamics [25, 26].

*(C) sequence of TauR2. [Source: Bhandare et al, 2019; doi: 10.1038/s41598-019-47249-7].*

*DOI: http://dx.doi.org/10.5772/intechopen.95792*

*Homology Modeling of Tubulin Isotypes to Investigate MT-Tau Interactions DOI: http://dx.doi.org/10.5772/intechopen.95792*

**Figure 1.**

*Homology Molecular Modeling - Perspectives and Applications*

this challenging problem in the field of Alzheimer's disease.

and other neurodegenerative diseases associated with tau [11, 12].

Full-length structure of tau protein is not yet determined using X-ray crystallographic techniques due to its intrinsically disordered nature. Also, the efforts to find its solution structure using NMR spectroscopy have failed [13]. Thus, the MT-Tau interactions have been studied so far using various biochemical and biophysical techniques [14–17]. The CryoEM have showed marginal success in determining the structure of tau repeat R2 bound to MT however it shows discontinues density of tau repeats along with each protofilament upon MT binding [8]. Hence, they synthetically developed R1 and R2 repeat of tau and their interactions with MT were examined. These two tau repeats adopts the extended conformation along the crest of protofilament which stabilizes the MT structure by binding to the interface

MTs are made from αβ-tubulin heterodimer subunits [18]. In human, seven α-tubulin and nine β-tubulin isotypes are reported showing their tissue-specific expressions. For instance, βI tubulin isotype is ubiquitously expressed in all cells,

detailed knowledge on MT-Tau interactions are still lacking mainly because of two reasons (i) Lack of full-length structure of tau due to its intrinsically disordered nature and (ii) Differential expression of tubulin isotypes in different type of cells in particular brain and neuronal cells. Earlier experimental efforts have been made to elucidate these interactions using solution structure (PDB ID: 2MZ7.pdb) however correct binding mode and atomistic interactions at structural level are poorly understood. Therefore, this chapter focuses on application of molecular modeling techniques in understanding important MT-Tau interactions in the Alzheimers disease. Bioinformatics approaches like sequence analysis, homology modeling. MD simulations and binding energy calculations are employed systematically to address

Tau is intrinsically disordered protein encoded by '*mapt'* gene located on chromosome 17 [2]. The primary function of the tau protein is to bind and stabilize the microtubule. It is abundantly expressed in the brain and neuronal tissues hence its misregulation is associated with the Alzheimers and other neurodegenerative disorders [3, 4]. Till date about six isoforms of tau are reported in the human central nervous system. The length of these six isoforms varies between 352 to 441

Primary structure of tau contains the projectile domain at N-terminal (residue 1–244) which is composed of the acidic and proline-rich region, and the C-terminal repeat domain which consists of 4 repeats i.e., R1, R2, R3 and R4 (residues 245–441) (**Figure 1**). The six isoforms of tau mainly differs by the existence of either R3 or R4 repeats at the C-terminal domain [6]. The one of the tau isoforms is referred as longest isoform mostly observed in humans which comprises 4 repeats i.e., R1, R2, R3 and R4. while the shortest isoform of tau has only 3 repeats (R1, R2 and R3). This shortest isoform of tau is reported in the fetus brain and less common in adults [5, 7]. **Figure 1A** represents the structure of tau repeat region R2 which is bound to the MT composed of β/α/β tubulin subunits [8]. Hereafter, tau repeat R2 will be mentioned as 'TauR2' for the simplicity. The tau repeats R1, R2, R3 and R4 prefers to bind at the outer surface of microtubule (MT) to stabilize it (**Figure 1A**) and regulates MT polymerization [6]. **Figure 1B** represents domain organization in the tau primary structure and **Figure 1C** shows the sequence of TauR2 which is reported in the CryoEM model. It is well established that tau primarily helps in the assembly and stabilization of axonal MTs, which contributes to the proper functioning of neuronal cells [9]. However, recent studies have reported new functional role of the tau in addition to the axonal, i.e. labile domain of the MT to promote its assembly [10]. Tau detaches from the MTs and forms abnormal, fibrillar structures of insoluble aggregates due to post-translational modifications in Alzheimer diseases

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of tubulin dimers [8].

residues [5].

*CryoEM Structure of tubulin subunits bound to TauR2. (A) tubulin subunits bound to TauR2 in CryoEM structure 6CVN.pdb. TauR2 domain binds at the outer surface of the MT. (B) Domain organization in tau, (C) sequence of TauR2. [Source: Bhandare et al, 2019; doi: 10.1038/s41598-019-47249-7].*

βII and βIII tubulin isotype are mainly expressed in brain and neuronal cells, and βVI tubulin isotype is expressed in erythroid cells and platelets [19]. The βI tubulin isotype reported to play crucial role in cell viability, βII tubulin isotype is important for neurite growth and βIII tubulin isotype protects nerve cell against free radicals and reactive oxygen species [20]. It has been well known that all β-tubulin isotypes share a significant residue conservation except the C-terminal tail region of MT [21–24] which is flexible in nature and structurally disordered. The C- tail region of all these isotypes overhang outwards of the MTs. The C-tail shows interactions with various MAPs including tau and regulate MT dynamics [25, 26].

It is well documented that the composition of β-tubulin isotypes (i) affects MT dynamic instability [27, 28], (ii) their interaction with motor proteins [29], (iii) their binding to the anti-drugs [21, 22, 30] and (iv) different MAPs including tau [31, 32]. These tubulin isotypes show tissue specific expression as their relative proportion varies greatly in different type of cells [20, 33, 34]. It is also well established that binding of tau to the MT promote or demote microtubule polymerization [35]. However, the differential binding affinity of tau to the various β-tubulin isotypes

expressed in different types of cells is completely unknown. Therefore, we studied relative binding affinity of Tau repeat region R2 with neuronal specific β-tubulin isotypes namely βI, βIIb, and βIII using molecular modeling [36].
