The calpain system originally comprised three molecules: two Ca2+-dependent proteases, mu-calpain and m-calpain, and a third polypeptide, calpastatin, whose only known function is to inhibit the two calpains. Both mu- and m-calpain are heterodimers containing an identical 28-kDa subunit and an 80-kDa subunit that shares 55-65% sequence homology between the two proteases. The crystallographic structure of m-calpain reveals six "domains" in the 80-kDa subunit: 1). a 19-amino acid NH2-terminal sequence; 2). and 3). two domains that constitute the active site, IIa and IIb; 4). domain III; 5). an 18-amino acid extended sequence linking domain III to domain IV; and 6). domain IV, which resembles the penta EF-hand family of polypeptides. The single calpastatin gene can produce eight or more calpastatin polypeptides ranging from 17 to 85 kDa by use of different promoters and alternative splicing events. The physiological significance of these different calpastatins is unclear, although all bind to three different places on the calpain molecule; binding to at least two of the sites is Ca2+ dependent. Since 1989, cDNA cloning has identified 12 additional mRNAs in mammals that encode polypeptides homologous to domains IIa and IIb of the 80-kDa subunit of mu- and m-calpain, and calpain-like mRNAs have been identified in other organisms. The molecules encoded by these mRNAs have not been isolated, so little is known about their properties. How calpain activity is regulated in cells is still unclear, but the calpains ostensibly participate in a variety of cellular processes including remodeling of cytoskeletal/membrane attachments, different signal transduction pathways, and apoptosis. Deregulated calpain activity following loss of Ca2+ homeostasis results in tissue damage in response to events such as myocardial infarcts, stroke, and brain trauma.

The Calpain System

Molecular self-assembly is the spontaneous association of molecules under equilibrium conditions into stable, structurally well-defined aggregates joined by noncovalent bonds. Molecular self-assembly is ubiquitous in biological systems and underlies the formation of a wide variety of complex biological structures. Understanding self-assembly and the associated noncovalent interactions that connect complementary interacting molecular surfaces in biological aggregates is a central concern in structural biochemistry. Self-assembly is also emerging as a new strategy in chemical synthesis, with the potential of generating nonbiological structures with dimensions of 1 to 10(2) nanometers (with molecular weights of 10(4) to 10(10) daltons). Structures in the upper part of this range of sizes are presently inaccessible through chemical synthesis, and the ability to prepare them would open a route to structures comparable in size (and perhaps complementary in function) to those that can be prepared by microlithography and other techniques of microfabrication.

Molecular self-assembly and nanochemistry: A chemical strategy for the synthesis of nanostructures

There is a special reason for reviewing this book at this time: it is the 50th edition of a compendium that is known and used frequently in most chemical and physical laboratories in many parts of the world. Surely, a publication that has been published for 56 years, withstanding the vagaries of science in this century, must have had something to offer. There is another reason: while the book is a standard fixture in most chemical and physical laboratories, including those in medical centers, it is not as frequently seen in the laboratories of physician's offices (those either in solo or group practice). I believe that the Handbook can be useful in those laboratories. One of the reasons, among others, is that the various basic items of information it offers may be helpful in new tests, either physical or chemical, which are continuously being published. The basic information may relate

Handbook of Chemistry and Physics

Hutchinson-Gilford progeria syndrome (HGPS) is a rare genetic disorder characterized by features reminiscent of marked premature ageing. Here, we present evidence of mutations in lamin A (LMNA) as the cause of this disorder. The HGPS gene was initially localized to chromosome 1q by observing two cases of uniparental isodisomy of 1q-the inheritance of both copies of this material from one parent-and one case with a 6-megabase paternal interstitial deletion. Sequencing of LMNA, located in this interval and previously implicated in several other heritable disorders, revealed that 18 out of 20 classical cases of HGPS harboured an identical de novo (that is, newly arisen and not inherited) single-base substitution, G608G(GGC > GGT), within exon 11. One additional case was identified with a different substitution within the same codon. Both of these mutations result in activation of a cryptic splice site within exon 11, resulting in production of a protein product that deletes 50 amino acids near the carboxy terminus. Immunofluorescence of HGPS fibroblasts with antibodies directed against lamin A revealed that many cells show visible abnormalities of the nuclear membrane. The discovery of the molecular basis of this disease may shed light on the general phenomenon of human ageing.

/pdf/recurrent-de-novo-point-mutations-in-lamin-a-cause-1kpya5najd.pdf

Recurrent de novo point mutations in lamin A cause Hutchinson-Gilford progeria syndrome

https://mmbr.asm.org/content/mmbr/47/1/1.full.pdf

Comparison of initiation of protein synthesis in procaryotes, eucaryotes, and organelles.

https://febs.onlinelibrary.wiley.com/doi/pdf/10.1016/0014-5793%2889%2981584-4

Maturation of nuclear lamin A involves a specific carboxy‐terminal trimming, which removes the polyisoprenylation site from the precursor; implications for the structure of the nuclear lamina

The intermediate filament proteins vimentin, desmin, and glial fibrillary acidic protein are cleaved in vitro by human immunodeficiency virus type 1 protease (HIV-1 PR). Microsequencing showed that HIV-1 PR cleaved both human and murine vimentin between leucine-422 and arginine-423 within the sequence between positions 418 and 427, Ser-Ser-Leu-Asn-Leu/Arg-Glu-Thr-Asn-Leu (SSLNL/RETNL). Minor cleavages at other sites were also observed. Heat-denatured vimentin was cleaved by HIV-1 PR less efficiently than native vimentin. A decapeptide containing the sequence SSLN-LRETNL was also cleaved in vitro by HIV-1 PR as predicted. The presence of a charged residue (arginine) at the primary cleavage site distinguishes this from other known naturally occurring cleavage sites. Microinjection of HIV-1 PR into cultured human fibroblasts resulted in a 9-fold increase in the percentage of cells with an altered and abnormal distribution of vimentin intermediate filaments. Most commonly, the intermediate filaments collapsed into a clump with a juxtanuclear localization. These results support the possibility that intermediate filament proteins may serve as substrates within HIV-1-infected cells.

https://www.pnas.org/content/pnas/87/16/6336.full.pdf

Human immunodeficiency virus type 1 protease cleaves the intermediate filament proteins vimentin, desmin, and glial fibrillary acidic protein.

The in vitro DNA-binding properties of purified nuclear lamin proteins and vimentin.

Solid-phase binding assays with protein species purified from cultured rat glioma C6 cells and Ehrlich ascites revealed that plectin bound specifically to lamin B but not to lamins A and C. Lamin B interaction was significantly decreased upon in vitro phosphorylation of either lamin B or plectin with protein kinase A or C. In contrast, phosphorylation of plectin with kinase A increased its binding to vimentin, suggesting a different regulation of plectin interactions by this kinase. 32P-radiolabeling of rat glioma C6 cells revealed plectin as a major in vivo target of protein kinase A and protein kinase C. Plectin, present in lysates of dibutyryladenosine 3',5'-cyclic monophosphate-treated cells, showed a 2.5 times higher binding affinity to vimentin than plectin from phorbol ester-treated cells. Furthermore, the relative amounts of plectin in 1% Triton X-100/high salt-insoluble cell fractions decreased to one-fourth of control values upon treating cells with phorbol esters, whereas vimentin was unaffected. This finding suggested a protein kinase C-dependent weakening of plectin interaction with intermediate filaments in vivo. Taken together, these results point to a role of plectin in interlinking cytoskeletal and nuclear elements and suggest that specific protein kinases are involved in regulating these interactions.

Protein kinase A- and protein kinase C-regulated interaction of plectin with lamin B and vimentin.

A Ca2+-activated neutral protease is described which, when tested against various native proteins, appears to be specific for vimentin, the 58,000-Mr subunit protein of intermediate-sized (7--11 nm) filaments in Ehrlich-ascites-tumour cells. The protein subunits of other classes of intermediate-sized filaments have been tested; neurofilament protein and glial fibrillary acidic protein are not degraded, however skeletin, the subunit protein of intermediate-sized filaments in smooth muscle, is degraded. The protease is found associated with the detergent-resistant cytoskeleton of Ehrlich-ascites-tumour cells; proteins, other than vimentin, present in this structure are not degraded. The protease is activated by Ca2+ and Sr2+ but not by other divalent cations tested: the Ca2+ concentration required for activation is 10 microM. The pH optimum is between pH 7.5 and 8.0, and the KCl concentration required for optimal activity is 100 mM. The protease is inhibited by 1-chloro-3-tosylamido-7-amino-L-2-heptanone hydrochloride and L-1-tosylamido-2-phenylethyl chloromethyl ketone but not by soybean trypsin inhibitor; inhibition by phenylmethylsulphonyl fluoride is moderate. The high substrate specificity of the protease suggests it may play a role in vimentin intermediate-sized filament protein turnover in Ehrlich-ascites-tumour cells.

https://febs.onlinelibrary.wiley.com/doi/pdf/10.1111/j.1432-1033.1981.tb05299.x

Peter Traub

Papers

Maturation of nuclear lamin A involves a specific carboxy‐terminal trimming, which removes the polyisoprenylation site from the precursor; implications for the structure of the nuclear lamina

Human immunodeficiency virus type 1 protease cleaves the intermediate filament proteins vimentin, desmin, and glial fibrillary acidic protein.

The in vitro DNA-binding properties of purified nuclear lamin proteins and vimentin.

Protein kinase A- and protein kinase C-regulated interaction of plectin with lamin B and vimentin.

Properties of a Ca2+-Activated Protease Specific for the Intermediate-Sized Filament Protein Vimentin in Ehrlich-Ascites-Tumour Cells