ArtsAutosBooksBusinessEducationEntertainmentFamilyFashionFoodGamesGenderHealthHolidaysHomeHubPagesPersonal FinancePetsPoliticsReligionSportsTechnologyTravel

The properties of mammalian phosphofructokinase (PFK)

Updated on November 5, 2011

Phosphofructokinase (PFK) is the enzyme responsible for catalysing the irreversible third reaction in glycolysis [2] which involves the addition of a phosphate group to fructose 6-phosphate (F6P) creating fructose 1,6-bisphosphate (FbisP) (Figure 1). Glycolysis is the metabolic pathway that breaks down glucose into pyruvate releasing free energy which is stored as ATP (adenosine triphosphate) and NADH (nicotinamide adenine dinucleotide). In response to conditions inside and outside of the cell the rate of glycolysis is tightly controlled and may even be reversed (glucogenisis). PFK has a central role in the regulation of glycolytic flux.

Figure 1 Reaction 3 of glycolysis in which fructose 6-phosphate is converted to fructose 1,6-bisphosphate by the addition of a phosphate group. The reaction is catalysed by PFK
Figure 1 Reaction 3 of glycolysis in which fructose 6-phosphate is converted to fructose 1,6-bisphosphate by the addition of a phosphate group. The reaction is catalysed by PFK

To synthesise PFK the DNA sequence must be transcribed onto mRNA which is then passed out of the nucleus for translation by the ribosomes. As a cytosolic enzyme PFK is likely to be produced by free ribosomes which aggregate amino acids together into a polypeptide chain. The sequence of the amino acids is known as the primary structure and is unique to a particular protein[1].

The chain of amino acid residues does not remain in a linear form but folds into a precise higher-order structure. The form this takes is largely derived from the order of the amino acids and a range of non-covalent interactions between them. The chain will, in theory, assume the most ‘energetically favourable’ shape or conformation based on these interactions. These include the relative positioning of hydrophobic / hydrophilic residues and hydrogen bonds between the N-H and C=O groups. As a globular protein, PFK will be formed from a variety of non-covalently ordered regions (a-helices and ß-sheets) held together by loop regions. Whilst irregular this secondary structure is precise.

As a complex globular protein PFK has a correspondingly complicated tertiary structure, that is its three-dimensional form composed of several domains. These are functional, structurally independent sub-units held together by flexible hinge regions. Individual domains often form the shape of the binding site of the protein. PFK is a homotetrameric protein, meaning it is composed of four identical polypeptide chains held together by non-covalent bonding.

The precise three-dimensional structure of an enzyme’s active site is such that it complements the shape of it substrate. The exactness of this match will affect how substrate specific an enzyme is. Given that PFK an allosteric enzyme formed from four identical subunits there are some likely features of its structure. As mentioned a protein’s domains may act as functional units, e.g. forming the active site, either alone or in concert. In allosteric enzymes it is not unusual for the active site to exist between the domains of the separate polypeptide subunits. It may be that the allosteric properties come from a substrate or effector binding to one active site which causes a structural change on an other active site. This is the basis of both cooperativity and inhibition / activation.

PFK has another important role, it is perhaps the most important regulator of glycolytic flux. Like most regulatory enzymes PFK is an allosteric enzyme meaning its catalytic activity can be controlled by substances unrelated to its substrate [2]. Allosteric enzymes are particularly well adapted to responding to small fluctuations in concentrations of their own substrates and other effectors molecules, both of which can be seen in the functioning of PFK.

Figure 2 Plots of initial rate v against initial substrate concentration [S] for a) an allosteric enzyme and b) a non-allosteric enzyme
Figure 2 Plots of initial rate v against initial substrate concentration [S] for a) an allosteric enzyme and b) a non-allosteric enzyme

The kinetics of allosteric enzymes make them well suited to controlling the rate of reactions. Unlike other enzymes where the plot of initial reaction rate v against substrate concentration [S] is hyperbolic, allosteric enzymes produce a sigmoidal plot (Figure 2). This is due to the mechanism of cooperativity in which binding with the first substrate molecule makes it easier for the enzyme to bind with further substrate molecules. Hence on the plot we see a slow start followed by a rapid rise (Figure 2 b). The advantage of these kinetics is that they are highly responsive to changes in substrate concentrations over very small ranges, i.e. those that will be found in vivo.

PFK’s responsiveness to substrate concentrations is further enhanced as it catalyses a rate-determining step in glycolysis [2]. At cellular concentrations the ratio of PFK’s substrate to product are very far from equilibrium. Whilst the enzyme will function at a steady rate if the substrate to product ration remains constant it must compensate for any change. If the substrate/product levels are near equilibrium there is little scope for this.

PFK is an example of an enzyme showing feedback inhibition [1]. This commonly involves an enzyme early in a metabolic pathway being inhibited by the products of a later enzyme-catalysed reaction in the pathway. This happens with PFK in resting skeletal muscle and what is of note here is that the pathway continues by being fed an alternative fuel at a later stage.

Figure 3 Regulation of glucose oxidation in skeletal muscle showing the roles and effectors of allosteric proteins (in bold)
Figure 3 Regulation of glucose oxidation in skeletal muscle showing the roles and effectors of allosteric proteins (in bold)

When at rest skeletal muscle oxidises triacylglycerols (TAG) as opposed to glucose which is prioritised for the brain. TAGs can be broken down into fatty acids and glycerol and fed into the glucose oxidation pathway as alternative fuels. Glycerol is converted into DHAP and fatty acids enter the tricarboxylic (TCA) cycle as acetyl CoA. It is the product of the first reaction in the TCA cycle (T1), citrate, that inhibits PFK causing a build up in its substrate, F6P. This in turn causes the second reaction (G2) of glycolysis to be reversed leading to a build up of glucose 6-phosphate (G6P). This build up of G6P prevents glycogen being broken down into glucose as it acts as an inhibitor to glycogen phosporylase, the enzyme responsible for catalysing the break down. The only remaining source of glucose is from the blood and update of this is prevented by fatty acids directly inhibiting the GLUT4 glucose transporters in the cell membrane.

A further way in which PFK regulates glycolytic flux is in how it responds to AMP (adenosine monophosphate) concentrations within the cell. In order to maintain a ready supply of ATP the enzyme adenylate kinase catalyses the conversion of ADP into ATP and AMP:

The concentration of AMP in cells is very low and whilst the above reaction has little effect on the relative concentration of ATP it has a significant effect on AMP concentration. AMP is a powerful allosteric activator of PFK so the effect is to increase glycolytic flux and rapidly restore the ATP levels.

Whilst not everything about the functioning of enzymes is clearly understood, as proteins a great deal is known about their precise molecular structure and synthesis. PFK plays a crucial role as a catalyst in what is perhaps the most fundamental of all metabolic pathways, glycolysis. What is of particular interest is how PFK is central to the regulation of glycolysis. Through both feedback inhibition and activator effectors PFK activity can be regulated. Its effectiveness and responsiveness are further enhanced by cooperativity and the far from equilibrium concentrations of its substrate / product. The position of PFK close to the beginning of the glycolytic pathway further establishes it as the major regulator of glycolytic flux.

References

  1. Walker C. & Swithenby M. (2007) ‘Proteins: Structure and Catalytic Function’ in Jill Saffery (ed) The Core of Life. S204 Book 3 Volume I 2nd edn. The Open University
  2. Furth A. (2007) ‘Metabolism’ in Jill Saffery (ed) The Core of Life. S204 Book 3 Volume I 2nd edn. The Open University

Word count: 1,146 words

This essay was from a second level biology course with the Open University and I think it scored mid-nineties.

Comments

    0 of 8192 characters used
    Post Comment

    No comments yet.

    working

    This website uses cookies

    As a user in the EEA, your approval is needed on a few things. To provide a better website experience, hubpages.com uses cookies (and other similar technologies) and may collect, process, and share personal data. Please choose which areas of our service you consent to our doing so.

    For more information on managing or withdrawing consents and how we handle data, visit our Privacy Policy at: https://hubpages.com/privacy-policy#gdpr

    Show Details
    Necessary
    HubPages Device IDThis is used to identify particular browsers or devices when the access the service, and is used for security reasons.
    LoginThis is necessary to sign in to the HubPages Service.
    Google RecaptchaThis is used to prevent bots and spam. (Privacy Policy)
    AkismetThis is used to detect comment spam. (Privacy Policy)
    HubPages Google AnalyticsThis is used to provide data on traffic to our website, all personally identifyable data is anonymized. (Privacy Policy)
    HubPages Traffic PixelThis is used to collect data on traffic to articles and other pages on our site. Unless you are signed in to a HubPages account, all personally identifiable information is anonymized.
    Amazon Web ServicesThis is a cloud services platform that we used to host our service. (Privacy Policy)
    CloudflareThis is a cloud CDN service that we use to efficiently deliver files required for our service to operate such as javascript, cascading style sheets, images, and videos. (Privacy Policy)
    Google Hosted LibrariesJavascript software libraries such as jQuery are loaded at endpoints on the googleapis.com or gstatic.com domains, for performance and efficiency reasons. (Privacy Policy)
    Features
    Google Custom SearchThis is feature allows you to search the site. (Privacy Policy)
    Google MapsSome articles have Google Maps embedded in them. (Privacy Policy)
    Google ChartsThis is used to display charts and graphs on articles and the author center. (Privacy Policy)
    Google AdSense Host APIThis service allows you to sign up for or associate a Google AdSense account with HubPages, so that you can earn money from ads on your articles. No data is shared unless you engage with this feature. (Privacy Policy)
    Google YouTubeSome articles have YouTube videos embedded in them. (Privacy Policy)
    VimeoSome articles have Vimeo videos embedded in them. (Privacy Policy)
    PaypalThis is used for a registered author who enrolls in the HubPages Earnings program and requests to be paid via PayPal. No data is shared with Paypal unless you engage with this feature. (Privacy Policy)
    Facebook LoginYou can use this to streamline signing up for, or signing in to your Hubpages account. No data is shared with Facebook unless you engage with this feature. (Privacy Policy)
    MavenThis supports the Maven widget and search functionality. (Privacy Policy)
    Marketing
    Google AdSenseThis is an ad network. (Privacy Policy)
    Google DoubleClickGoogle provides ad serving technology and runs an ad network. (Privacy Policy)
    Index ExchangeThis is an ad network. (Privacy Policy)
    SovrnThis is an ad network. (Privacy Policy)
    Facebook AdsThis is an ad network. (Privacy Policy)
    Amazon Unified Ad MarketplaceThis is an ad network. (Privacy Policy)
    AppNexusThis is an ad network. (Privacy Policy)
    OpenxThis is an ad network. (Privacy Policy)
    Rubicon ProjectThis is an ad network. (Privacy Policy)
    TripleLiftThis is an ad network. (Privacy Policy)
    Say MediaWe partner with Say Media to deliver ad campaigns on our sites. (Privacy Policy)
    Remarketing PixelsWe may use remarketing pixels from advertising networks such as Google AdWords, Bing Ads, and Facebook in order to advertise the HubPages Service to people that have visited our sites.
    Conversion Tracking PixelsWe may use conversion tracking pixels from advertising networks such as Google AdWords, Bing Ads, and Facebook in order to identify when an advertisement has successfully resulted in the desired action, such as signing up for the HubPages Service or publishing an article on the HubPages Service.
    Statistics
    Author Google AnalyticsThis is used to provide traffic data and reports to the authors of articles on the HubPages Service. (Privacy Policy)
    ComscoreComScore is a media measurement and analytics company providing marketing data and analytics to enterprises, media and advertising agencies, and publishers. Non-consent will result in ComScore only processing obfuscated personal data. (Privacy Policy)
    Amazon Tracking PixelSome articles display amazon products as part of the Amazon Affiliate program, this pixel provides traffic statistics for those products (Privacy Policy)