Introduction

Figure 1.

Lipoprotein lipase (LPL) is an enzyme synthesized and secreted primarily by myocytes and adipocytes. 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 the diglyceride and monoglyceride products 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 can have detrimental effects on

GPIHBP1 deficiency or mutations that affect GPIHBP1 disulfide bond formation is a cause of chylomicronemia, an inability to break down lipids, since GPIHBP1 no longer has the capacity to bind to LPL.^[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. PBD file to 6OB0 LPL complexed with GPIHBP1

LPL

LPL is composed of an and a .

N-terminal Domain

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 the lid region which is in an open conformation, meaning it is not blocking the active site. The consists of 2 short α-helices connected by a loop. ^[1]

C-terminal Domain

The of LPL includes the GPIHBP1 binding site and the that helps to contribute to the specificity of this enzyme for TRLs. ^[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. Within this hydrophobic region is the access point to the .

Mechanism

The active site consists of a catalytic triad SER-159, ASP-183 and HIS-268 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 monoglyceride, is released.

^[3]

^[1]

^[2]

^[5]

Inhibitor

Medical Relevance

Mutations

Loss of function mutations in LPL or GPIHBP1 cause a number of genetic diseases, the most prominent being hypertriglyceridemia, or high levels of triglycerides in the blood. Two specific point mutations that have been directly related to 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.

Additionally, the mutation p.D201V 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. This, in turn, destabilizes LPL folding and prevents its secretion from cells.