The High Resolution Crystal Structure of the Human Tumor Suppressor Maspin Reveals a Novel Conformational Switch in the G-helix

Abstract

Maspin is a serpin that acts as a tumor suppressor in a range of human cancers, including tumors of the breast and lung. Maspin is crucial for development, because homozygous loss of the gene is lethal; however, the precise physiological role of the molecule is unclear. To gain insight into the function of human maspin, we have determined its crystal structure in two similar, but nonisomorphous crystal forms, to 2.1-and 2.8-Å resolution, respectively. The structure reveals that maspin adopts the native serpin fold in which the reactive center loop is expelled fully from the A ␤-sheet, makes minimal contacts with the core of the molecule, and exhibits a high degree of flexibility. A buried salt bridge unique to maspin orthologues causes an unusual bulge in the region around the D and E ␣-helices, an area of the molecule demonstrated in other serpins to be important for cofactor recognition. Strikingly, the structural data reveal that maspin is able to undergo conformational change in and around the G ␣-helix, switching between an open and a closed form. This change dictates the electrostatic character of a putative cofactor binding surface and highlights this region as a likely determinant of maspin function. The high resolution crystal structure of maspin provides a detailed molecular framework to elucidate the mechanism of function of this important tumor suppressor.Maspin (mammary serine proteinase inhibitor (SERPINB5)) was initially identified as a tumor-suppressing serpin downregulated in invasive mammary carcinoma cell lines (1). Maspin loss in numerous cancers (including breast, prostate, squamous cell carcinoma, gastric cancer, and lung) correlates with metastasis and a poor clinical prognosis (for a review, see Ref.2). In contrast, high levels of maspin expression in certain cancers (in particular, pancreatic and ovarian cancer) correlate with tumor invasion and poor survival. Like other clade B serpins (3), maspin has a nucleocytoplasmic distribution, however it is also found at the cell surface (1, 4, 5). The intracellular role of maspin is at present unclear, but it has been suggested to play a role in apoptosis pathways (6). A large body of evidence suggests that maspin has an important extracellular role: it can suppress tumor growth and metastasis in vivo and tumor cell motility and invasion in vitro (1,7,8). Maspin also plays a fundamental role in early embryonic development; murine knock-out studies reveal that it is essential for proper organization of the epiblast (9). Consistent with a complex role in tumorigenesis, maspin also exhibits anti-angiogenic activity (10), and expression of the maspin gene has been demonstrated to be under the control of the oncogenic transcription factors p53 and p63 (11,12). Because the failure to properly control proteolytic activity can result in disruption of the basement membrane and promote tumor invasion, it was initially hypothesized that maspin may exert its anti-metastatic effect by functioning as a pericellular protease inhibitor (1).Serpins are unusual ...

G-helix of Maspin Mediates Effects on Cell Migration and Adhesion

“…Although early evidence suggested that maspin was an inhibitory serpin able to block plasminogen activation by urokinase plasminogen activator and tissue-type plasminogen activator (11)(12)(13), we demonstrated that this was not the case in a number of conditions where the serpin PAI-1 was inhibitory (9). That maspin is a noninhibitory serpin is supported by crystal structure data revealing that its RCL does not correspond with those found in inhibitory serpins (14,15). It remains possible that maspin influences protease activity indirectly by noninhibitory interactions with the plasminogen activators (16,17) and protection of matrix from degradation by cathepsin D (18).…”

Endogenous Inhibition of Histone Deacetylase 1 by Tumor-Suppressive Maspin

“…However, X-ray crystallographic analyses of maspin revealed that maspin has a general serpin conformation with an exposed reactive site loop and an novel G-helix that may allow a significant amount of flexibility for induced conformational changes (13). This suboptimal serpin conformation may confer a distinct preference for partners, such as pro-uPA (32), that are not entirely committed to a serine protease conformation.…”

“…Based on its high-resolution crystal structure, maspin is indeed a noninhibitory serpin against active serine proteases. Nonetheless, maspin retains the basic serpin conformation with an exposed reactive site loop (12,13). Thus, maspin may interact with serine protease-like partners or other proteins.…”

“…Thus, maspin may interact with serine protease-like partners or other proteins. The binding of maspin to a serine protease-like molecule and other proteins may be uniquely accommodated by its conformational heterogeneity of the G-helix and its novel unstable shutter region salt bridge (13). In the meantime, breakthroughs in decoding the maspin mechanisms came from advances in the identification of diverse molecular partners/targets of maspin, including tissue-type plasminogen activator (14), pro-urokinase type plasminogen activator (pro-uPA; ref.…”

“…A major breakthrough in translating the sequence code of maspin came from X-ray crystallographic analyses of the maspin structure (12,13). Based on its high-resolution crystal structure, maspin is indeed a noninhibitory serpin against active serine proteases.…”