Yama/CPP32β, a mammalian homolog of CED-3, is a CrmA-inhibitable protease that cleaves the death substrate poly(ADP-ribose) polymerase

Caspases, which are the executioners of apoptosis, comprise two distinct classes, the initiators and the effectors. Although general structural features are shared between the initiator and the effector caspases, their activation, inhibition and release of inhibition are differentially regulated. Biochemical and structural studies have led to important advances in understanding the underlying molecular mechanisms of caspase regulation. This article reviews these latest advances and describes our present understanding of caspase regulation during apoptosis.

Molecular mechanisms of caspase regulation during apoptosis

Heparin, a sulfated polysaccharide belonging to the family of glycosaminoglycans, has numerous important biological activities, associated with its interaction with diverse proteins. Heparin is widely used as an anticoagulant drug based on its ability to accelerate the rate at which antithrombin inhibits serine proteases in the blood coagulation cascade. Heparin and the structurally related heparan sulfate are complex linear polymers comprised of a mixture of chains of different length, having variable sequences. Heparan sulfate is ubiquitously distributed on the surfaces of animal cells and in the extracellular matrix. It also mediates various physiologic and pathophysiologic processes. Difficulties in evaluating the role of heparin and heparan sulfate in vivo may be partly ascribed to ignorance of the detailed structure and sequence of these polysaccharides. In addition, the understanding of carbohydrate-protein interactions has lagged behind that of the more thoroughly studied protein-protein and protein-nucleic acid interactions. The recent extensive studies on the structural, kinetic, and thermodynamic aspects of the protein binding of heparin and heparan sulfate have led to an improved understanding of heparin-protein interactions. A high degree of specificity could be identified in many of these interactions. An understanding of these interactions at the molecular level is of fundamental importance in the design of new highly specific therapeutic agents. This review focuses on aspects of heparin structure and conformation, which are important for its interactions with proteins. It also describes the interaction of heparin and heparan sulfate with selected families of heparin-binding proteins.

http://www-heparin.rpi.edu/main/files/papers/4f1623c54d6ab5.21304086.pdf

Heparin-protein interactions

Almost one-third of all proteases can be classified as serine proteases, named for the nucleophilic Ser residue at the active site. This mechanistic class was originally distinguished by the presence of the AspHis-Ser “charge relay” system or “catalytic triad”.1 The Asp-His-Ser triad can be found in at least four different structural contexts, indicating that this catalytic machinery has evolved on at least four separate occasions.2 These four clans of serine proteases are typified by chymotrypsin, subtilisin, carboxypeptidase Y, and Clp protease (MEROPS nomenclature;3 Table 1). More recently, serine proteases with novel catalytic triads and dyads have been discovered, including Ser-His-Glu, Ser-Lys/His, His-Ser-His, and N-terminal Ser.2 Several of these novel serine proteases are subjects of accompanying articles in this issue. This article will review recent work on the mechanism and specificity of chymotrypsin-like enzymes, with the occasional references to pertinent experiments with subtilisin. Chymotrypsin-like proteases are the most abundant in nature, with over 240 proteases recognized in the MEROPS database.3 * E-mail: hedstrom@brandeis.edu; phone: 781-736-2333; FAX: 781-736-2349. 4501 Chem. Rev. 2002, 102, 4501−4523

Serine protease mechanism and specificity

The American Heart Association, in conjunction with the National Institutes of Health, annually reports the most up-to-date statistics related to heart disease, stroke, and cardiovascular risk factors, including core health behaviors (smoking, physical activity, diet, and weight) and health factors (cholesterol, blood pressure, and glucose control) that contribute to cardiovascular health. The Statistical Update presents the latest data on a range of major clinical heart and circulatory disease conditions (including stroke, congenital heart disease, rhythm disorders, subclinical atherosclerosis, coronary heart disease, heart failure, valvular disease, venous disease, and peripheral artery disease) and the associated outcomes (including quality of care, procedures, and economic costs).The American Heart Association, through its Statistics Committee, continuously monitors and evaluates sources of data on heart disease and stroke in the United States to provide the most current information available in the annual Statistical Update. The 2022 Statistical Update is the product of a full year's worth of effort by dedicated volunteer clinicians and scientists, committed government professionals, and American Heart Association staff members. This year's edition includes data on the monitoring and benefits of cardiovascular health in the population and an enhanced focus on social determinants of health, adverse pregnancy outcomes, vascular contributions to brain health, and the global burden of cardiovascular disease and healthy life expectancy.Each of the chapters in the Statistical Update focuses on a different topic related to heart disease and stroke statistics.The Statistical Update represents a critical resource for the lay public, policymakers, media professionals, clinicians, health care administrators, researchers, health advocates, and others seeking the best available data on these factors and conditions. 

https://www.ahajournals.org/doi/pdf/10.1161/CIR.0000000000001052

Heart Disease and Stroke Statistics—2022 Update: A Report From the American Heart Association

Antithrombin is unique among the serpins in that it circulates in a native conformation that is kinetically inactive toward its target proteinase, factor Xa. Activation occurs upon binding of a specific pentasaccharide sequence found in heparin that results in a rearrangement of the reactive center loop removing constraints on the active center P1 residue. We determined the crystal structure of an activated antithrombin variant, N135Q S380C-fluorescein (P14-fluorescein), in order to see how full activation is achieved in the absence of heparin and how the structural effects of the substitution in the hinge region are translated to the heparin binding region. The crystal structure resembles native antithrombin except in the hinge and heparin binding regions. The absence of global conformational change allows for identification of specific interactions, centered on Glu 381 (P13), that are responsible for maintenance of the solution equilibrium between the native and activated forms and establishes the existence of an electrostatic link between the hinge region and the heparin binding region. A revised model for the mechanism of the allosteric activation of antithrombin is proposed. Antithrombin is unique as a proteinase inhibitor because it requires allosteric activation. Its cofactor, heparin, binds antithrombin via a specific pentasaccharide domain that accounts for its anticoagulant activity. Antithrombin plays a critical role in the prevention of thrombosis by inhibiting the final two proteinases in the blood coagulation cascade, factor Xa and thrombin. The tight regulation of antithrombin involves the following: 1) maintenance of an inactive, native pool in plasma at 2.3 mM; 2) localization to the vascular endothelium through binding to glycosaminoglycans; 3) conformational activation; 4) ability to undergo a massive conformational rearrangement upon reaction with proteinase; and 5) release from glycosaminoglycan upon formation of complex with proteinase. The complexity of the role of antithrombin in hemostasis is illustrated by the wide range of antithrombin mutations that lead to

A 2.85-å structure of a fluorescein derivative reveals an electrostatic link between the hinge and heparin binding regions*

Structural insights unravel the zymogenic mechanism of the virulence factor gingipain K from Porphyromonas gingivalis , a causative agent of gum disease from the human oral microbiome.

Serpin polymerization is an event which generally occurs within living tissue as a consequence of a folding defect caused by point mutations. Major advances in cell biology and imaging have allowed detailed studies into subcellular localization, processing, and clearance of serpin polymers, but to understand the molecular basis of the misfolded state and polymeric linkage, it has been and continues to be necessary to generate polymers in vitro. The goal of this chapter is to outline the principal techniques that have been developed over the past 20 years to produce and characterize serpin polymerization in vitro. For the majority of this time, all data were interpreted in accordance with the so-called “loop-sheet” hypothesis, where polymers form through the intermolecular incorporation of the reactive center loop (RCL) of one serpin monomer into the β-sheet A of another. This hypothesis is supported by the ability of serpins to incorporate exogenous peptides into sheet A in an identical manner to the insertion of its own RCL upon cleavage by protease or conversion to the latent state. However, a recent crystal structure of an intact serpin dimer showed that much larger “domain swaps” are possible that would also lead to hyperstable linkage between serpin monomers. This chapter is therefore not limited to a description of experimental technique, but discusses the findings in light of the two current models of serpin polymerization. We would encourage readers to reevaluate the literature on serpin polymerization and to expand on the experiments outlined here in order to differentiate between possible domain-swapping mechanisms.

Serpin polymerization in vitro.

Thrombin is the ultimate coagulation factor. Not only is it the final protease generated by the blood coagulation cascade, it has more than 12 substrates and 5 cofactors. How thrombin specificity is directed during the four stages of hemostasis is of great interest to the medical community, as insufficient thrombin activity leads to bleeding and excessive activity results in thrombosis. Over the last three decades we have learned a great deal about how thrombin is generated and how it recognizes its several cofactors, substrates, and inhibitors. Although much has been inferred from biochemical studies, our current understanding is primarily based on numerous crystallographic structures of thrombin complexes. In this chapter I provide an overview of the multiple roles thrombin plays in the initiation, amplification, propagation, and attenuation phases of hemostasis, and describe how the special structural features of thrombin are exploited to achieve regulation and substrate selectivity.

Structural Insights into the Life History of Thrombin

https://febs.onlinelibrary.wiley.com/doi/pdf/10.1016/j.febslet.2006.07.057

James A. Huntington

Papers

A 2.85-å structure of a fluorescein derivative reveals an electrostatic link between the hinge and heparin binding regions*

Structural insights unravel the zymogenic mechanism of the virulence factor gingipain K from Porphyromonas gingivalis , a causative agent of gum disease from the human oral microbiome.

Serpin polymerization in vitro.

Structural Insights into the Life History of Thrombin

Allosteric activation of antithrombin is independent of charge neutralization or reversal in the heparin binding site.