Abstract
Cyclobenzaprine hydrochloride (CBH) is a well-known muscle relaxant that is widely used to relieve muscle spasms and other pain associated with acute musculoskeletal conditions. In this study, we elucidated the binding characteristics of this muscle relaxant to human serum albumin (HSA). From a pharmaceutical and biochemical viewpoint, insight into the structure, functions, dynamics, and features of HSA-CBH complex holds great importance. The binding of CBH with this major circulatory transport protein was studied using a combination of biophysical approaches such as UV-VIS absorption, fluorescence quenching, and circular dichroism (CD) spectroscopy. Various in silico techniques, molecular docking and molecular dynamics, were also used to gain deeper insight into the binding. A reduction in the fluorescence intensities of HSA-CBH complex with a constant increase in temperature, revealed the static mode of protein fluorescence quenching upon CBH addition, which confirmed the formation of the HSA-CBH ground state complex. The alteration in the UV-VIS and far-UV CD spectrum indicated changes in both secondary and tertiary structures of HSA upon binding of CBH, further proving CBH binding to HSA. The analysis of thermodynamic parameters ∆H° and ∆S° showed that binding of CBH to HSA was dominated by intermolecular hydrophobic forces. The results of the molecular docking and molecular dynamics simulation studies also confirmed the stability of the complex and supported the experimental results.
Highlights
Skeletal muscle, the largest organ of the human body, comprises 30–40% of the total body weight [1]
The findings show that in the presence of Cyclobenzaprine hydrochloride (CBH), there is a slight increase in the thermodynamic stability of human serum albumin (HSA), i.e., the presence of CBH increases the overall stability of HSA
The results obtained from circular dichroism (CD) showed that the binding of CBH leads to increased α-helical content of HSA
Summary
The largest organ of the human body, comprises 30–40% of the total body weight [1]. Cyclobenzaprine hydrochloride (CBH) (Figure 1), a 5-HT2 receptor antagonist, is a well-known muscle relaxant. The second derivative curves of absorption spectra (Figure 3) of HSA, in the absence and presence of CBH, allowed us to clearly monitor the changes occurring in the micro-. SSeeccoonndd ddeerriivvaattiivvee aabbssoorrppttiioonn ssppeeccttrraa ooff hhuummaann sseerruumm aallbbuummiinn ((HHSSAA)) iinn tthhee aabbsseennccee aanndd ppFriregesuseernnecc3ee.((2S20e0cμμoMnMd))odoffeCrCiBvBHaHtiavatetppaHHbs7o7..r44paatninoddnttseepmmepcpteerrraaatotuufrreheu22m55◦a°CnC..serum albumin (HSA) in the absence and presence (20 μM) of CBH at pH 7.4 and temperature 25 °C. Change in Gibbs free energy (ΔG°) for the HSA-CBH complex s2y.4s.teTmhearmt dodifyfenraemnitctPeamrapmereatetrusraensdisFsohrcoews nInivnolTvaedblien1H. Tehxpereerfiomree,ntths)e(pFoigsuitrivee5Bv)a.luTeheofsiΔgSn°s haenrdemisaagcnlietuardiensdoifcavtairoinouthsatht ehrymdorodpyhnoabmicicinptaerraamcteiotenrss asurechdoams tihneanent tirnotphyecbhianndgineg(∆oSf ◦C),BfHre-eHeSnAer.gAy nchegaantgivee(∆vGal◦u),eaonfdΔeHnt°hianldpiycactheasnbgoeth(∆hHy◦d)raorgeevnitbaol nindsanayndcovmapnledxerfoWrmaaatlsioanreanadsscoacniabteedutwiliitzhedthteo fiodremntaiftyiotnheofdtihffeerleignatntydp-persootfeifnorccoems ipnlveoxlv[2e9d].in the formation of HSA-CBH complex [27]. ∆S◦ here is a clear indication that hydrophobic interactions are dominant in the binding of CBH-HSA. A negative value of ∆H◦ indicates both hydrogen bonds and van der Waals are associated with the formation of the ligand-protein complex [29]
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