Download This Pdf To Understand The Classification Of Enzymes In Biochemistry

The EC classification groups enzymes that perform the same or related enzymatic functions. You can use the EC browser to explore enzymes that have similar functions, shape of enzyme and/or arrangements of amino acids performing the enzyme reaction. It can also be used to identify and explore structures of enzymes that have similar functions but different shapes and/or mechanism of enzyme reaction.

Classification Of Enzymes In Biochemistry Pdf Download

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There are three types of hydrogenase, the [NiFe], [FeFe] and [Fe] hydrogenases, that are distinguished by their metal composition. Whereas the [Fe]-hydrogenases are a small methanogenic-specific family12, the [NiFe] and [FeFe] classes are widely distributed and functionally diverse. They can be classified through a hierarchical system into different groups and subgroups/subtypes with distinct biochemical features (e.g. directionality, affinity, redox partners and localization) and physiological roles (i.e. respiration, fermentation, bifurcation, sensing)1,6. It is necessary to define the subgroup or subtype of the hydrogenase to predict hydrogenase function. For example, while Group 2a and 2b [NiFe]-hydrogenases share >35% sequence identity, they have distinct roles as respiratory uptake hydrogenases and H2 sensors respectively13,14. Likewise, discrimination between Group A1 and Group A3 [FeFe]-hydrogenases is necessary to distinguish fermentative and bifurcating enzymes2,15. Building on previous work16,17, we recently created a comprehensive hydrogenase classification scheme predictive of biological function2. This scheme was primarily based on the topology of phylogenetic trees built from the amino acid sequences of hydrogenase catalytic subunits/domains. It also factored in genetic organization, metal-binding motifs and functional information. This analysis identified 22 subgroups (within four groups) of [NiFe]-hydrogenases and six subtypes (within three groups) of [FeFe]-hydrogenases, each proposed to have unique physiological roles and contexts2.

We initially developed a classification scheme to enable prediction of hydrogenase function by primary sequence alone. To do this, we visualized the relationships between all hydrogenases in sequence similarity networks (SSN)18, in which nodes represent individual proteins and the distances between them reflect BLAST E-values. As reflected by our analysis of other protein superfamilies19,20, SSNs allow robust inference of sequence-structure-function relationships for large datasets without the problems associated with phylogenetic trees (e.g. long-branch attraction). Consistent with previous phylogenetic analyses2,16,17, this analysis showed the hydrogenase sequences clustered into eight major groups (Groups 1 to 4 [NiFe]-hydrogenases, Groups A to C [FeFe]-hydrogenases, [Fe]-hydrogenases), six of which separate into multiple functionally-distinct subgroups or subtypes at narrower logE filters (Fig. 1; Figure S1). The SSNs demonstrated that all [NiFe]-hydrogenase subgroups defined through phylogenetic trees in our previous work2 separated into distinct clusters, which is consistent with our evolutionary model that such hydrogenases diverged from a common ancestor to adopt multiple distinct functions2. The only exception were the Group A [FeFe]-hydrogenases, which, as previously-reported2,17, cannot be classified by sequence alone as they have principally diversified through changes in domain architecture and quaternary structure. It remains necessary to analyze the organization of the genes encoding these enzymes to determine their specific function, e.g. whether they serve fermentative or electron-bifurcating roles.

In addition to its precision, the classifier is superior to other approaches due to its usability. It is accessible as a free web service at HydDB allows the users to paste or upload sequences of hydrogenase catalytic subunit sequences in FASTA format and run the classification (Figure S2). When analysis has completed, results are presented in a table that can be downloaded as a CSV file (Figure S3). This provides an efficient and user-friendly way to classify hydrogenases, in contrast to the previous standard which requires visualization of phylogenetic trees derived from multiple sequence alignments30.

In addition to its classification function, HydDB is designed to be a definitive repository for hydrogenase retrieval and analysis. The database presently contains entries for 3248 hydrogenases, including their NCBI accession numbers, amino acid sequences, hydrogenase classes, taxonomic affiliations and predicted behavior (Figure S4). To enable easy exploration of the data set, the database also provides access to an interface for searching, filtering and sorting the data, as well as the capacity to download the results in CSV or FASTA format. There are individual pages for the 38 hydrogenase classes defined here (Table 1), including descriptions of their physiological role, genetic organization, taxonomic distribution and biochemical features. This is supplemented with a compendium of structural information about the hydrogenases, which is integrated with the Protein Databank (PDB), as well as a library of over 500 literature references (Figure S5).

According to the unified classification principle of enzymes published by the International Society of Biochemistry, each group of enzymes in the above seven categories can be further divided into several subgroups according to the characteristics of the functional groups or bonds in the substrates. In order to show the properties of substrates or reactants more accurately, each subclass is further divided into subclasses and directly contains a quantity of enzymes.

Mechanistically diverse superfamilies pose an especially difficult problem for automated functional classification methods due to the complexity of their underlying biology. For example, a newly sequenced superfamily member may not catalyze the same overall reaction as its closest relative in the superfamily, but may instead be related to other superfamily members by a more subtle conserved chemical capability. If the superfamily itself has not been characterized, the conserved chemical capability may not be immediately obvious. It is thus useful to subdivide a superfamily into families containing enzymes that catalyze the same overall reaction.

The main difference between our family classifications and those of Pfam and SCOP is their coverage of function space. As shown in Table 3, our gold and silver standard families include only sequences that catalyze a single overall reaction. Although some SCOP and Pfam families (for example, the enolase family) correspond to this level of functional similarity, Table 3 shows that most are broader, principally because these classification systems rely mainly on overall sequence and structural similarities rather than on the finer granularity analysis focused on the subsets of catalytic residues that distinguish enzymes that perform a specific catalytic reaction. For example, the Pfam MR_MLE_N and MR_MLE families include enzymes that catalyze at least seven different overall reactions. This difference is illustrated graphically in Figure 1.

Comparison of gold and silver standard family classifications to Pfam for the gold standard enolase superfamily. The outer ring represents Pfam family classifications. Sequences that match multiple Pfam HMMs, all of which correspond to a single SFLD functional domain (for example, 'Enolase_N', representing the amino terminus of the enzyme enolase and 'Enolase', representing the carboxyl terminus of the enzyme enolase), are shown with a single designation in the figure to simplify the illustration. (a) The inner ring represents gold standard family classifications. Gray regions represent enzymes that can be assigned to the gold standard enolase superfamily, but cannot be confidently assigned to a gold standard family. (b) The inner ring represents silver standard family classifications. Gray regions represent enzymes that can be assigned to the gold standard enolase superfamily, but cannot be confidently assigned to a silver standard family.

Figure 1 also shows that some of the enzymes in our gold standard enolase superfamily are classified into the Pfam IMPDH family, which contains inosine monophosphate dehydrogenases, among other enzymes. Although the members of the IMPDH family share the (β/α)8 (TIM) barrel fold common to enolase superfamily members, they do not have the amino-terminal domain found in all enolase superfamily members, nor do they use a similar set of catalytic residues to perform their functions. Thus, we believe that classification of any enolase superfamily members into the Pfam IMPDH superfamily is incorrect.

Superfamily classifications for four of our five gold standard superfamilies (amidohydrolase, enolase, haloacid dehalogenase, and vicinal oxygen chelate) correspond to the analogous SCOP and SUPERFAMILY superfamily designations. In contrast, the gold standard crotonase superfamily is only a subset of the corresponding Clp/crotonase superfamily in SCOP and SUPERFAMILY. The SCOP Crotonase-like family contains enzymes corresponding to the gold standard crotonase superfamily, while the remaining families listed in the SCOP Clp/crotonase superfamily contain enzymes that may be evolutionarily related to gold standard crotonase superfamily members, but do not have an established mechanistic linkage [42, 43]. Again, because there is no explicit indication of the functional similarity contained within a SCOP or SUPERFAMILY superfamily, it is difficult to use these classifications to make functional inferences regarding uncharacterized proteins.

In the development of the gold standard set, we encountered several difficulties in attempting to classify sequences that belong to mechanistically diverse superfamilies into their constituent families. These difficulties largely arise from the complexity of the underlying biology, where the boundaries between different families within a superfamily may be uneven due to different evolutionary rates within each family, and, due to a number of reasons, some enzymes may not fit into the simple family classification at all. 2ff7e9595c