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Biostatistics and Bioinformatics
Evolution of Enzymes
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Biostatistics and Bioinformatics Evolution of Enzymes
Description of Module Subject Name Biochemistry
Paper Name 13 Biostatistics and Bioinformatics Module Name/Title 20 Evolution of Enzymes
Dr. Vijaya Khader Dr. MC Varadaraj
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1. Objectives: The learning objectives for evolution of enzymes include
1.1. Compiling a set of orthologous enzymes with known kinetic values such as Km and Kcat, 1.2. Predicting consensus sequence of the enzyme
1.3. Predicting consensus secondary structure
1.4. Mapping the residues which are significant for improving performance of the enzymes, and Finally 1.5. Understanding gene duplication for creating paralogs with new activity
2. Concept Map
Paralogs Creating New Enzyme Activities Brief Description
Summary
OSBS MSA using PROMALS3D
Kinetic Values Dataset: OSBS
Compiling OSBS Sequence Dataset for MSA
Constructing MSA using PROMALS3D
Identifying Critical Residues
Conserved Catalytic Residues Conserved Binding Site Residues
Conserved Residues within Secondary Structures Conserved Prolines and Glycines
Conserved Residue in long Loop
SemiConserved and Poorly Conserved Residues
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3. Description
Dear students, we have seen in the last module, that during evolution, certain residue positions are conserved for motifs or single identities. In addition, we have seen that there are certain strong and weak similarities in positions of multiple sequence alignment. All enzymes are proteins, and therefore, we will, necessarily be learning the evolution of proteins in this module. Proteins are linear sequences of amino acids encoded in genes. The genes, after replication, are passed on to offsprings in next generation.
However, during replication, the gene sequence may undergo point mutations, both in its regulatory sequence to control the concentration of an enzyme within cell or may undergo mutations within protein coding sequence to affect the affinity or turnover number of an enzyme. Those individuals which acquire mutations suitable to survive in the changing environment may be selected as fittest i.e. the well known theory of Sir Charles Darwin i.e. the survival of the fittest. In module 13 and 14 we identified active site residues of OSBS from 3D structure alignment from substrate and product bound enzyme complexes.
Because 3D structure of the Hpr K/P bound with its substrate ATP or pyrophosphate is not available, therefore, in last module, we identified active site residues of Hpr K/P from multiple sequence alignment of closely and distantly related Hpr K/P sequences. This shows that both the structure and sequence approaches can be used to identify residues involved in catalysis. Therefore, in this module we will combine sequence and structure approaches for enhancing the performance of enzyme activity.
Km of an enzyme reflects its affinity for the substrate and low Km enables enzyme to work at low substrate concentration. During enzyme engineering, we may be interested in decreasing Km of an enzyme so as to enable it work at low substrate concentration. Similarly, Vmax of an enzyme, which is the product of Kcat and total enzyme concentration, reflects best performance in terms of turn over number. An enzyme with high turn over number is able to perform better at lower concentration of the enzyme. Therefore, enzyme industry may be interested in increasing turnover number of an enzyme, so that low concentration of the enzyme is required in the target operation. However, both Km and Kcat donot allow classifying enzymes on the basis of catalytic efficiency. Therefore, Kcat and Km are combined to get catalytic efficiency of the enzyme terms of ratio of Kcat and Km, i.e. Kcat/Km. Therefore, enzyme efficiency can be increased by either increasing Kcat or lowering Km. The Highest catalytic efficiency is defined that as soon as the substrate diffuses into the active site of an enzyme, it is transformed to its product without consuming further time.
Consequently, an enzyme may achieve its efficiency up to the upper limit of diffusion of substrate to active site, which is between 108 and 109 M-1s-1. This is exemplified by highly evolved enzymes such as acetylcholine esterase, triosephosphate isomerase, carbonic anhydrase and some others. However, the majority of enzymes, has achieved catalytic efficiency approximately one million M-1s-1.
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In addition to its main or major activity, sometimes an enzyme has some additional but minor activity or what we call a promiscuous activity. During replication, complete gene duplication may happen and duplicated copy of the gene may accumulate point mutations to convert its minor activity to main activity and thereby resulting in the evolution of a new enzyme. In this module, we will combine the consensus primary sequence with consensus secondary structure to identify point mutations for improving performance of an enzyme and we will look at gene duplications for acquiring new enzyme activity. The OSBS family has several members including both closely related and divergent amino acid sequences, as well as four 3D structures. Therefore OSBS provides an excellent model system, for elucidating principles of enzyme evolution.
Therefore, the learning objectives for understanding evolution of enzymes include compiling a set of orthologous enzymes with known kinetic values such as Km and Kcat. Then we need to predict consensus sequence of the enzyme which determines the kinetic parameters of the enzyme. We also need to predict consensus secondary structure which is mapped on to the predicted consensus sequences for correct folding of the enzyme. This can be achieved with multiple sequence alignment application available at PROMALS3D. Then, we will map the residues which are significant for improving performance of the enzymes. Finally, we look at gene duplication for creating paralogs with new activity. In this module we will understand evolution of enzymes to identify significant residues which can be useful for enhancing performance of Enzymes and gene duplications for creating new activities.
3.1. Multiple Sequence Alignment of OSBS using PROMALS3D
PROMALS3D allows prediction of consensus secondary structure which is mapped on to the predicted consensus sequences. Therefore, we will use Promals3D for constructing OSBS multiple sequence alignment.
3.1.1. Compiling OSBS Kinetic Values Dataset
Ortho-succinylbenzoate synthase (OSBS) belongs to the enolase superfamily of enzymes, characterized by the presence of an enolate intermediate, which is generated by abstraction of the alpha-proton of the carboxylate substrate.
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OSBS catalyses the dehydration reaction of SHCHC to produce product o-succinylbenzoate. In the modules 13 and 14, we took the example of enzyme OSBS, i.e. o-succinylbenzoate synthase, to search PDB database and found enzyme structures bound to its substrate and product and could detect active site residues from these complexes. In this module, we will carry forward the same example to identify other essential residues for enhancing performance of the enzyme.
Visit enzyme database at ExPASy available at http://enzyme.expasy.org/ and search for o-succinylbenzoate synthase.
The search displays enzyme 4.2.1.113 for o-succinylbenzoate synthase with several alternative names.
Visit BRENDA enzyme kinetics database and search for 4.2.1.113 and press start search button.
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The EC Number 4.2.1.113 is the required enzyme.
Therefore, click this enzyme and in the subsequent page Limit your search to E. coli
and expand functional parameters.
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Km data from E. coli reveals that there are several mutations which result in both increase and decrease in Km i.e. affinity of OSBS for its substrate. However, most of the mutations lead to increase in Km i.e.
reduced affinity from 0.012 milimolar for wildtype enzyme. This can be understood simply because these mutations were planned in laboratory and the mutated OSBS is not selected under environmental pressure for survival of the fittest enzyme with highest affinity. But, nevertheless this data will help us understanding point mutations for altering affinity, and consequently, substrate specificity. For example, mutation of serine at 262, which is detected in binding site of enzyme-substrate 3D complex, results in increased Km. Mutation to threonine results in drastic increase in Km as opposed to slight increase in mutation to glycine. The mutation at I265A increases affinity by a factor of 10. Similarly, Kcat data from E.
coli reveals that there are several mutations which result in both increase and decrease in Kcat i.e.
turnover number of OSBS for its substrate.
There are several mutations which lead to increase in Kcat i.e. increased turnover number. This data will help us understanding point mutations for altering turnover number and consequently, Vmax. In addition, different mutations at the same position may have different effects on Kcat value. For example mutation
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of serine at 262, to glycine results in decreased Kcat whereas, mutation to threonine enhances Kcat.
Kcat/Km list for E. coli reveals that wild type enzyme has catalytic efficiency i.e. Kcat/Km at 2000 mM-1s-1 (2
* 106 M-1s-1) and the substitution of Serine at 262 with glycine reduces the catalytic efficiency by one fourth, whereas substitution with threonine at same position results in to one tenth. The OSBS from E. coli has catalytic efficiency of 2 million M-1s-1. On the other hand, OSBS from Thermobifida fusca has catalytic efficiency of 0.67 million M-1s-1. Therefore, OSBS with catalytic efficiency on either side of one million M-1s-1 and 80 closely and distantly related members, is an excellent model system for identifying critical residues for improving enzyme efficiency. But before identifying the amino acid residues affecting the performance of OSBS from multiple sequence alignment, let us check if Kcat and Km data values are available for mutated OSBS analogs.
This clearly reveals the involvement of the serine 262 in the binding of the substrate as the catalytic efficiency is not affected drastically. Kcat/Km list for E. coli reveals the wild type enzyme is mutated twice at position 51. In addition mutant at position 51 has another mutation at position 48. The position 48 was also mutated individually, to see effect of mutation at position 48 only. The substitution of phenylalanine at 51 with alanine reduces the efficiency to 400 whereas substitution with tyrosine at same position results in reduction to 350. However, second mutation at L48M in F51Y increase efficiency to 370, whereas single mutation at L48M with 570 is better than even double mutant with 370. From these results it is clear that all mutations reduced catalytic efficiency of wild type from 2000 mM-1s-1 (2 * 106 M-1s-1). These catalytic efficiencies reveal that all mutant enzymes have resulted in unfavourable catalytic efficiency.
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The periodic spectrum of amino acids shows that the mutations at positions 48, 51, 262, 288 were planned as per physicochemical properties.
For example, replacement of leucine with methionine at 48 or phenylalanine with tyrosine at 51, or serine with threonine at 262 or glycine with alanine at 288, were introduced keeping physicochemical properties
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in the periodic spectrum of amino acids. Therefore, mutating any of these residues as per physico-chemical similarities is expected to either increase or decrease catalytic efficiency, slightly. However, drastic reduction in Kcat/Km from 2000 to 3.8 indicates that only physico-chemical criteria for planning mutations are not sufficient. Therefore, let us check if the secondary structure at these positions is also important for evolving catalytic efficiency of enzymes. For this purpose, we need consensus sequence and Consensus secondary structure, which can be predicted using multiple sequence alignment using Promals3D application. For constructing MSA, we need closely and distantly related OSBS amino acid sequences.
Go to Concept Map
3.1.2. Compiling OSBS Sequence Dataset for MSA
Visit UniProtKB and search for o-succinylbenzoate synthase, Limit search to reviewed 82 and select 250 from Show dropdown list to display all 82 sequences in single page. Now Sort the entries on the basis of length. When we sort on the basis of length, then an option to restrict range of length option is presented below tool bar. Scrolling entries shows that first two entries with unusual length of 1715 and 451 do not belong to OSBS. The next entries with length 371 to last entry with length 320 are all OSBS.
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Therefore, limit the search to length from 320 to 371, by clicking on the restrict range of length and entering this range.
Now select all and download 80 selected sequences in uncompressed format. The entries will appear in the browser window.
Copy on clipboard and paste in NotePad.
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Find the entry for entry P29208, the entry from E. coli, copy it and paste at the end of file. Save the file with a file name having extension name FASTA and ‘save as type’ all files.
Go to Concept Map
3.1.3. Constructing MSA using PROMALS3D
PROMALS3D available at prodata.swmed.edu/promals3d/promals3d.php, is a profile based multiple sequence alignment application which combine PSIPRED to predict secondary structure for constructing multiple sequence alignment. Promals3D first aligns similar sequences and then uses PSI-BLAST and PSIPRED results for aligning divergent sequences. Promals3D applies more elaborate techniques to align relatively divergent clusters. It allows extraction of sequences from PDB structure files also.
Therefore upload the saved sequences in file for 80 OSBS sequences in FASTA format and upload 1FHU OSBS structure to click Submit Button. This will begin multiple sequence alignment and will update the progress every 30 seconds. When the alignment is complete, display colored alignment in the order of input sequences with 60 residues in each row and displaying residues with consensus level at 0.8.
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Therefore, in each column the consensus for the residues to the level of 0.8 i.e. 80% will be evaluated before displaying the residue in a consensus category.
The multiple sequence alignment with PROMALS3D returns the consensus amino acid sequence in the second last line of alignment block and consensus predicted secondary structure in the last line. The secondary structure elements are represented with symbol “h” for alpha helices and symbol “e” for extended beta strands. The positions without any of these symbols are either turns or loops. Identical amino acids (with chosen consensus level, 0.8 in the present example) are displayed in uppercase and bold letters. The other consensus amino acid positions are categorized under categories. I will come back to these categories in a little while from now.
Go to Concept Map
3.2. Identifying point mutations for Improving Enzyme Efficiency
The absolute conservation within or at the ends of secondary structure revealed the presence of catalytic residues. We know from modules 13 and 14 that lysine at position 133 is a catalytic base for abstraction of alpha proton from carboxylate substrate. Similarly, the aspartate at 161, glutamate at 190 and aspartate at 213 are involved in coordination of magnesium ion to stabilize alpha carboxyl in transition state. The arginine at 235 is involved in stabilization of enolate ion position in the transition state.
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3.2.1. Conserved Catalytic Residues
The Lys133, Lys235 and ASP161, GLU190, ASP213 are the residues involved in chemical catalysis.
Therefore, mutating any of these catalytic residues is expected to abolish activity completely. The PROMALS3D found these five residues conserved. Therefore, if a mutation at any conserved residue abolish the activity completely, that means the residue is involved in chemical mechanism. Consequently, PROMALS3D MSA can be used for identification of catalytic residues in enzymes for which the catalytic residues are not identified.
The mapped active site for OSBS shows that the Mg2+ ion is coordinated by ASP161, GLU190, ASP213 and one oxygen of the carboxylate group of OSB.
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The literature information also shows that Lys(133) and Lys(235) are positioned to function as acid/base catalysts in the dehydration reaction of substrate to produce OSB. Both these residues are position on each face of the planar product OSB.
3.2.2. Conserved Binding Site Residues
The MSA found S262 conserved at the end of beta stand at position 262.
Therefore, mutation with threonine or glycine results in decrease in catalytic efficiency of OSBS. The bound product in 3D complex with OSBS reveals the presence of serine 262 inside the binding site for positioning the substrate.
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However, surprisingly, mutation with threonine, although strong similarity, yet it results in 10 fold decrease in catalytic efficiency. Whereas, replacement with a similar tiny amino acid glycine at this position at the end of secondary structure still allows some comfort with reducing efficiency by a factor of 4 to 480. This shows the absolute requirement for conserved serine as both increasing size with preserving hydroxyl and decreasing size without hydroxyl, reduce the catalytic efficiency. However, the reduction in catalytic efficiency by just a factor of 10 indicates the involvement of position in binding rather than in catalysis.
Therefore, PROMALS3D MSA can be used for identification of binding residues in enzymes for which the binding residues are not identified. However, mutating serine 262 is expected to affect catalytic efficiency, but not drastically, as serine 262 is not involved in catalysis. The 3D structure of OSBS bound to OSB shows presence of hydroxyl group of serine near to carboxylate group of succinyl moiety of OSB. Therefore, this increases affinity of enzyme for OSB.
For mutating a residue in its 3D structure, open SwissPDBViewer and open PDB structure 1FHV containing OSBS bound to OSB. Display the residue to mutate, Ser262 in the present case and select Mutate tool, the second last tool from Tool bar.
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It will ask to pick a group, choose Ser262 by clicking over its alpha carbon and from ensuing context menu chose THR fro mutating to threonine and click Mutate tool again and respond by accepting mutation sSer262 to Thr262. This will mutate the residue Ser262 to Thr262.
Replacing serine with threonine preserves hydroxyl for affinity (shown with hydrogen bond displayed in dotted green line) but introduces methyl which clash with carboxylate group of succinyl moiety of OSB (shown with pink color clash line from carboxylate of OSB to methyl group in Threonine) , thereby reducing catalytic efficiency.
Therefore, predicted conservation at 262 indicates that this position is significant and shall not be mutated for improving enzyme catalytic efficiency. The mutation with glycine leaves open space to reduce catalytic efficiency. Therefore, predicted conservation at 262 indicates that this position is significant and shall not be mutated for improving enzyme catalytic efficiency.
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3.2.3. Conserved Prolines and Glycines
The conserved residues glycine126, proline154 and glycine240 are present within short stretches. The conserved proline within short stretches at the end of a regular secondary structure is required for introducing a turn to change direction of secondary structure advancement for correct 3D folding.
Similarly, conserved glycine within a short stretch is mostly involved in turning a loop. Therefore, predicted conservations at 126, 154 and 240 indicate that this position is significant and shall not be mutated as it will deteriorate catalytic efficiency. The predictions for the presence of glycine126, proline154 and glycine240 short stretches reveals introduction of a turn or a small loop in 3D structure as found in available 3D structure of E. coli OSBS displayed in red colored turns and loops.
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3.2.4. Conserved Residue in long Loop
A conserved glycine 288 is present at almost within a loop of 11 residues. Available kinetic data for mutations at these position reveals drastic reduction in kinetic efficiency from 2000 M-1s-1 to 3.8 M-1s-1. The drastic reduction in kinetic efficiency of mutations at this conserved position is expected. Let us now have a look at the reason for the drastic reduction in kinetic efficiency of mutations at this conserved position within a long loop.
Replacement of glycine with alanine at position 288 reduces catalytic efficiency by a factor approximately 500. From 3D structure, it is clear that mutation from glycine to alanine at position 288 results in the clash of the introduced methyl group in alanine with carboxyl moiety of succinyl group in the substrate.
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Therefore, substrate is not able to fit in the binding pocket correctly, thereby reducing the catalytic efficiency.
3.2.5. Conserved Residues within Secondary Structures
Let us have a look at the presence of conserved residues within long secondary structures. Conserved arginine at 27 is present at almost within middle of a long beta strand of 14 residues, and another conserved arginine at 159 is present exactly at middle of a beta strand of 5 residues. Available kinetic data for mutation at position 159 reveals drastic reduction in kinetic efficiency from 2000 M-1s-1 to 11 M-1s-1. The drastic reduction in kinetic efficiency of mutations at this conserved position is expected.
Let us now have a look at the reason for the drastic reduction in kinetic efficiency of mutations at this conserved position within long secondary structures A conserved arginine at 159 is present exactly with a
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conserved pattern of hydrophobicity with spacing 2 having intervening variable hydrophilic residues. This spacing with variable hydrophilic residues suggests a beta strand on the surface. The conservation of a arginine in beta strand on the surface indicate interaction for stabilization of 3D structure through electrostatic interactions. Replacement of arginine with methionine in E. coli reduces catalytic efficiency by a factor 200. This suggests the involvement of arginine 159 in folding of the enzyme. The presence of arginine in E. coli OSBS position at 159 in 3D structure reveals its electrostatic interactions with main chain carbonyl from Isoleucine 286 connecting two regular secondary structures separated by more than 100 amino acids in the primary sequence.
We donot have the kinetic data for mutation at conserved arginine at 27, present at almost within middle of a long beta strand of 14 residues. Therefore, let us see if this arginine is involved in electrostatic interactions for connecting two regular secondary structures separated by a long stretch of amino acids in the primary sequence. In SwissPDBViewer, choose Arg27, choose “Compute H-bonds” command from tool menu, then from display menu choose “Show Only H-bonds from selection and the choose “Show Only Groups with Visible H-bond” and display complete chain as ribbons from fifth column of control panel. It is clear from 3D structure that conserved arginine at 27 has several electrostatic interactions with carbonyl groups of main atoms of distantly placed S264, I265, S46 and P47. Therefore, mutati ng this residue will result in drastic decrease in the catalytic efficiency of the enzyme.
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3.2.6. SemiConserved and Poorly Conserved Residues
In figure next, all aligned positions are displayed.
Identical amino acids (with chosen consensus level, 0.8 in the present example) are displayed in consensus sequence in uppercase and bold letters. For example for the first 60 aligned positions, we have tyrosine, proline, arginine, two glycines and then glutamate. The non-identical consensus amino acid positions are categorized under 11 categories and displayed using separate symbols. The presence of hydroxyl group
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containing serine or threonine is indicated by lower case “o” letter. The negatively charged residues aspartate and glutamate is indicated with “-” sign and positively charged residues lysine, arginine or histidine is indicated with “+” sign. The lower case “l “ in italics indicate presence of any of three small aliphatic amino acids valine, leucine or Isoleucine. The conservation of the any one of the four tiny amino acids glycine, alanine serine and cysteine is indicated my lower case “t” letter. Sometimes, more liberal mutations are allowed at particular positions. For example, any one of the nine polar amino residues (D/E/H/K/N/Q/R/S/T) present in an aligned column is indicated with lower case “p” letter. This group adds four residues i.e. N/Q/S/T to five charged amino acids. There may be any one of the nine small amino acids (A, G, C, S, V, N, D, T, P) indicated with lower case “s” letter. Therefore more than five residues may be present in those columns. The conservation of the any one of the five charged amino acid i.e. H/R/K/D/E in a column is indicated with lower case “c” letter. Any one of ten bulky residues (E/F/I/K/L/M/Q/R/W/Y) indicated with lower case “b” letter, or any on of the eleven hydrophobic residues (W/F/Y/M/L/I/V/A/C/T/H) indicated with lower case “h” letter. The “@” symbol indicate the presence of any one of the four aromatic amino acids phenylalanine, tryptophan, tyrosine or histidine. These categories indicate the reservation of the one of the 11 types of the amino acid groups in a particular column, which actually represent the requirement of the type of residue at the corresponding 3D position in structure. Therefore, positions with residues, especially for first five groups, falling within loops indicate mutations for improving enzyme performance. The mutation at I265A increases affinity for the substrate by a factor of 10. This is a mutation categorized under group four for aliphatic.
Therefore, multiple sequence alignment consensus sequence with consensus secondary structure provides insights into detecting significant functional residues in enzymes.
Go to Concept Map
3.3. Paralogs Creating New Enzyme Activities
Now let us have look at another aspect of enzyme evolution, the complete gene duplication creating additional copy of the same gene. Two situations may arise. First is if the original gene has some other activity, may be minor, also known as promiscuous activity, then this additional copy may undergo point mutations to convert this minor activity into the main activity and thereby creating new enzyme adding enhanced capability to the harboring individual. We know that enzyme OSBS has minor epimerase activity.
We will take example of OSBS i.e. a synthase with promiscuous epimerase activity for gene duplication and then analyzing point mutations to evolve this promiscuous activity into a separate epimerase for enhancing capabilities. Secondly, gene duplication may enhance performance of an individual /species by evolving Cooperativity among subunits as exemplified by hemoglobin chain paralogs.
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Let us check if duplicated OSBS gene with promiscuous Epimerase activity can evolve into main enzyme with epimerase activity and add new enzyme activity for evolving the harboring individual with additional capability. Visit UniProtKB at uniprot.org and search for L-Ala D/L Glu Epimerase. The search returns six sequences. Download these six sequences.
Paste these sequences at the beginning of already saved FASTA file with 80 sequences and save with a new file name, such as 80 OSBS and 6 Epimerase Sequences.FASTA.
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Upload saved file 80 OSBS and 6 Epimerase Sequences.FASTA at PROMALS3D available at prodata.swmed.edu/promals3d/promals3d.php and submit for multiple sequence alignment.
This will begin multiple sequence alignment and will update the progress every 30 seconds. When the alignment is complete, display colored alignment in the order of input sequences with 60 residues in each row and displaying residues with consensus level at 0.8. Therefore, in each column the consensus for the residues to the level of 0.8 i.e. 80% will be evaluated before displaying the residue in a consensus category.
It was clear from 3D structure in OSBS that conserved arginine at 27 has several electrostatic interactions with carbonyl groups of main atoms of distantly placed S264, I265, S46 and P47 for placing catalytic residues in correct 3D position to catalyze OSBS reaction. The arginine at this position has been replace with A, S, T, I or V in six epimerase sequences indicating loss of electrostatic interactions to fold epimerase differently for positioning the same catalytic residues to catalyze epimerase reaction.
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The presence of arginine in E. coli OSBS position at 159 reveals its electrostatic interactions with main chain carbonyl from Isoleucine 286 connecting two regular secondary structures separated by more than 100 amino acids in the primary sequence. The replacement of this arginine by Isoleucine or alanine in three epimerase enzymes again shows difference in folding pattern for positioning the same catalytic residues to catalyze different chemical reaction.
Replacement of glycine with alanine at position 288 reduces catalytic efficiency by a factor approximately 500. From 3D structure, it is clear that mutation from glycine to alanine at position 288 results in the clash of the introduced methyl group in alanine with carboxyl moiety of succinyl group in the substrate.
Similarly, the replacement of glycine at this position with aspartate in each of the six enzymes is responsible to reduce main OSBS activity and convert main activity into promiscuous OSBS activity.
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We know OSBS is main activity and epimerase is promiscuous activity. During evolution, promiscuous epimerase activity was transformed into main epimerase activity. Therefore, this confirms the role of mutations affecting folding pattern and substrate binding into creation of new activities.
Therefore, conversion of epimerase to main synthase could be achieved with single mutation from aspartate to glycine at position 288, as aspartate at 288 will not able to block the orientation of substrate
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for synthase activity. But the catalytic efficiency will be much low as several other mutations from synthase to epimerase ahs also occurred.
Go to Concept Map 4. Summary
Dear students, in this module we have seen the evolution of enzymes for improving existing activity for surviving under changing cellular environments. For example, the point mutations in OSBS for catalyzing same chemical reactions in Enteric and lactic acid bacteria allows selection of the fittest to function under changed environment from GI tract to lactic acid fermentation. Similarly, we have seen gene duplication and subsequent point mutations for optimizing the new activity. In this way, the nature allows evolution of new species to survive under different environments with enhanced capabilities. I thank you all for visiting ePGPathshala.
