Enzyme Promiscuity: Engine of Evolutionary Innovation

Abstract

Catalytic promiscuity and substrate ambiguity are keys to evolvability, which in turn is pivotal to the successful acquisition of novel biological functions. Action on multiple substrates (substrate ambiguity) can be harnessed for performance of functions in the cell that supersede catalysis of a single metabolite. These functions include proofreading, scavenging of nutrients, removal of antimetabolites, balancing of metabolite pools, and establishing system redundancy. In this review, we present examples of enzymes that perform these cellular roles by leveraging substrate ambiguity and then present the structural features that support both specificity and ambiguity. We focus on the phosphatases of the haloalkanoate dehalogenase superfamily and the thioesterases of the hotdog fold superfamily.In the 1990s, a series of studies on the evolution of catalysis in protein fold families helped define contemporary understanding of enzymes as potentially promiscuous catalysts; the analyses of these enzyme superfamilies suggested that certain folds showed higher variability than expected with regard to the chemistries that can be catalyzed or the substrates that can be acted on (1-11). To summarize, the current model holds that enzyme families grow as a result of gene duplication coupled with the acquisition of an advantageous new function. Because the backbone folds, and thus, the catalytic scaffolds are inherited, so is the chemical trait that underlies the intrinsic catalytic functions of all family members. In enzyme families, evidence can be found for low level intrinsic activity associated with one or more extant members, co-existing with the high level of activity unique to the subject enzyme (see for instance, the enolase and alkaline phosphatase enzyme superfamilies (12, 13)). The ability to carry out such alternate chemistry is termed catalytic promiscuity. The plausible link between catalytic promiscuity and evolvability has been explored in previous publications (for recent coverage and reviews of this topic, see Refs. 14 -17).The most commonly encountered observation of promiscuity involves the catalysis of one type of chemistry with many different substrates. Jensen (18) referred to this trait as "substrate ambiguity," and this is the name we will use. Herein, we examine the selective advantage associated with activity toward multiple substrates by highlighting specific examples of enzymes for which the level of substrate ambiguity runs high to fulfill specific roles in the cell. We use as examples enzymes from the haloalkanoate dehalogenase (HAD) 3 superfamily and the thioesterases of the hotdog fold superfamily. In addition, we dissect the architectures of enzymes from these families to discover underlying structural sources of specificity and substrate ambiguity. Screening to Assess Substrate AmbiguityIn vitro enzyme activity measurements carried out with a structurally diverse library of potential substrates allow one to generate a substrate specificity profile for the enzyme of interest. However, the mo...

Functional Diversity of Haloacid Dehalogenase Superfamily Phosphatases from Saccharomyces cerevisiae

“…2). Previously, pronounced substrate promiscuity has been demonstrated for HAD phosphatases from E. coli and other organisms (28,32,40). This broad substrate specificity apparently contributes to the key role of these enzymes in the hydrolysis of a broad range of phosphomonoesters as well as the "house-cleaning" functions.…”

“…This broad substrate specificity apparently contributes to the key role of these enzymes in the hydrolysis of a broad range of phosphomonoesters as well as the "house-cleaning" functions. Enzyme substrate promiscuity (also known as substrate ambiguity) is likely to be the starting point for the evolution of new enzymes through gene duplication followed by subfunctionalization (substrate specialization) of the diverging enzymes (28,127). Beneficial promiscuous activities in the promiscuous repertoire can be selected and improved through mutations, initially without losing the primary enzyme activity (127).…”

“…This superfamily was originally named after the haloacid dehalogenases, but it is dominated by putative phosphatases (ϳ79%) and ATPases (ϳ20%) and also includes phosphonatases and phosphomutases (25)(26)(27). Specifically, the HAD-like phosphatases are a family of diverse enzymes that are responsible for the majority of metabolic phosphomonoester hydrolysis reactions in all kingdoms of life (26,28). Extensive sequence comparisons show that the HAD-like hydrolases are defined by the presence of four conserved sequence motifs with the characteristic N-terminal motif I containing two Asp residues (DXD), whereas motifs II and III contain a highly conserved Thr (or Ser) and Lys, respectively (24,25).…”

A Computational Biology Approach in Function Annotation to Enzymes

“…This ability to perform alternate chemistry is known as catalytic promiscuity. The most common type of promiscuity is the ability of enzyme to catalyze one chemical reaction with different substrates, referred to as "substrate ambiguity" (Pandya et al, 2014). Besides the phenomenon of substrate ambiguity, the promiscuity can include other, different phenomena, depending on the circumstances.…”

“…In terms of function, it can be advantageous to the cell through several mechanisms such as 1) proofreading, 2) scavenging of nutrients, 3) removal of antimetabolites, 3) balancing of metabolite pools, and 4) establishing system redundancy (Pandya et al, 2014). When enzymes are classified according to six main classes, and EC numbers that are related to functions, their distribution in main classes and subclasses are as in Table 1, as of February 2018.…”