Introduction

Figure 1. LPL catalyzes the breakdown of triglycerides into a monoglyceride and two fatty acids.

Lipoprotein lipase (LPL) is an enzyme synthesized and secreted primarily by myocytes and adipocytes into interstitial spaces. It is located on the surface of capillaries where it is bound to a glycolipid-anchored protein expressed by capillary endothelial cells. This protein is called glycosylphosphatidylinositol-anchored high density lipoprotein-binding protein 1, or GPIHBP1. LPL is an essential enzyme for triglyceride metabolism and utilization, however it is susceptible to unfolding in its catalytic domain and thus must be bound to GPIHBP1 to prevent loss of enzymatic activity. When LPL is not bound to GPIHBP1 its enzymatic activity is relatively low and declines until it has lost all function, but when bound to GPIHBP1 it is able to maintain its maximum enzymatic activity.^[1] In addition, binding to GPIHBP1 is required for adhesion of triglyceride rich lipoproteins (TRLs) to LPL and transport of LPL to its site of action in the capillary lumen. Once it has reached the site of action the enzyme is able to produce a monoglyceride and two fatty acids from the triglyceride substrate (Figure. 1).

Function

Energy is metabolized through the hydrolysis of triglyceride-rich lipoproteins (TRLs), among other macromolecules, to release fatty acids that can be used or stored for later energy use ^[2]. LPL is the main enzyme involved with the metabolism of triglycerides within TRLs. The LPL-GPIHBP1 complex is crucial for clearing triglycerides from the bloodstream and delivery of lipid nutrients to vital areas, such as heart and skeletal muscle, and adipose tissue ^[3]. Loss of function mutations in LPL or GPIHBP1 can have detrimental effects on the body's ability to metabolize triglycerides which in turn causes chylomicronemia. Elevated levels of plasma triglycerides, called hypertriglyceridemia, also increases the risk of developing coronary artery disease (CAD)^[4].

Structure

The asymmetric unit is a tetramer of LPL/GPIHBP1 complexes. The orientation of these four dimers are in a . Despite the unique orientation of each of the four dimers within the tetramer, all act independently and perform the same enzymatic function.

[[Image:surfacerep.png|200px|right|thumb|Figure 2. A surface representation of LPL complexed with GPIHBP1.]

N-terminal α/β-hydrolase domain of LPL

The contains the by which this enzyme catalyzes the hydrolysis of triglycerides, these residues are Ser159, Asp183, and His268, and also houses the to stabilize the transition state of the substrate through the backbone amides of Trp82 and Leu160. The N-terminal domain includes a which has been shown to have mutations that may impact LPL enzyme activity. The lid region of the N-terminal domain was imaged in an open conformation, meaning it is not blocking the active site. The consists of 2 short α-helices connected by a loop, extending away from the protein. This open conformation allows for many surface-exposed hydrophobic residues (valines,isoleucines, and leucines) to create a hydrophobic patch on the surface of LPL. The lid region helps to control for the entry of lipid substrates into the active site cleft.^[1]

C-terminal β-barrel domain of LPL

The of LPL includes the GPIHBP1 binding site and the that helps to contribute to the specificity of this enzyme for TRLs by creating a second hydrophobic patch on the same face of LPL as the active site. The hydrophobic patches are believed to allow for the enzyme to bind TRL substrates with an orientation that facilitates delivery of triglycerides into the active site for catalysis. ^[1]

GPIHBP1

GPIHBP1 is a membrane bound protein that binds to the C-terminal domain of LPL. It contains specific features, the LU domain and the acidic domain, that contribute to its binding affinity. GPIHBP1’s LU domain is 75 residues in length and adopts a that is stabilized by . The LPL-GPIHBP1 binding interface involves the three-fingered fold and depends largely on hydrophobic interactions between the two subunits. It contains a highly acidic and disordered N-terminal domain with 21 of the 26 total residues being aspartates or glutamates which were not visible in the electron density map. This indicates that the acidic domain of GPIHBP1 likely exhibits conformational flexibility.^[3]

Active Site

The active site, located in the N-terminal domain is surrounded by a that stabilizes the hydrophobic lipid substrate in the binding pocket and forms van der Waals interaction with its hydrophobic tails. Within this hydrophobic region is the access point to the .

Mechanism

The active site consists of a catalytic triad Ser159, Asp183 and His268 that go through the serine protease mechanism of action. The hallmark of this mechanism is the proton shuttle between the three residues that increases the nucleophilicity of the serine residue. Serine is then able to make the nucleophilic attack on the the carbonyl carbon of the scissile peptide bond of the substrate. During catalysis, an ordered mechanism occurs in which several intermediates are generated. The catalysis of the peptide cleavage can be seen as a ping-pong catalysis, in which the triglyceride binds, and the diglyceride is released. Then the second substrate, water, binds and the second product, the fatty acid, is released.

Inhibitor

Medical Relevance

Mutations

Missense mutations in LPL or GPIHBP1 cause a number of genetic diseases, the most prominent being Chylomicronemia, or the inability to break down lipids. Two specific point mutations that contribute to this disease through the abolishment of LPL-GPIHBP1 binding are p.M404R and p.C445Y. Because the LPL-GPIHBP1 complex is bound together primarily by hydrophobic interactions, switching a nonpolar methionine residue to a positively charged arginine residue, or a nonpolar cysteine residue to a polar tyrosine residue will disrupt these hydrophobic interactions and weaken the interface. Also, the much larger side chain of arginine is difficult to accommodate into the hydrophobic pocket of GPIHBP1 ^[3]. These mutations do not affect LPL’s activity or secretion but affect its ability to be transported to its site of action in the capillary lumen. This leads to an accumulation of unmetabolized lipids but also catalytically active LPL in the interstitial spaces^[5].

Additionally, observed in patients with Chylomicronemia has been observed to eliminate LPL secretion and reduce its activity. Among other residues, the carboxylic acid side chain of D201 is critical for coordinating LPL’s calcium ion. Mutating this negatively charged aspartic acid into a nonpolar valine residue disrupts the ionic bond between calcium and the aspartate, disturbing the overall calcium binding. A similar mutation has also been observed for p.D202E. These two mutations, in turn, destabilize LPL folding and thereby prevent its secretion from cells^[3].